IS6110-based restriction fragment length polymorphism (RFLP) analysis of Mycobacterium tuberculosis H37Rv and H37Ra.

IS6110-based restriction fragment length polymorphism (RFLP) analysis of Mycobacterium tuberculosis H37Rv and its avirulent mutant H37Ra was performed by a number of restriction enzymes, including Nru I, EcoN I, Pst I, and Pvu II. No differences were found in the IS 6110-fingerprints of the study strains by Nru I. One differential IS6110-positive restriction fragment was detected by EcoN I in strain H37Ra, while analysis by Pst I revealed that two fragments of the strain H37Rv were replaced by four novel IS6110-positive fragments in the strain H37Ra. By using Pvu II, a restriction enzyme that cleaves IS 6110 once, and by probing for an IS6110 specific target sequence located to the right of the Pvu II site, we found that the strains H37Rv and H37Ra share 13 IS6110-positive restriction fragments and that one IS6110-positive restriction fragment of H37Rv is replaced by four novel fragments in H37Ra; by probing for an IS6110-specific target sequence to the left of the Pvu II site, 13 shared restriction fragments and 2 differential bands in strain H37Ra were detected. These findings demonstrate that novel insertions of the IS6110 element exist in the avirulent strain H37Ra and raise the question of the role, if any, of IS6110-insertional mutagenesis in the establishment of the avirulent M. tuberculosis H37Ra phenotype.

The antiviral activity of a synthetic peptide derived from the envelope SU glycoprotein of feline immunodeficiency virus maps in correspondence of an amphipathic helical segment.

In a previous paper (Lombardi et al., Virology 220, 274-284, 1996), we-reported that a 20-amino acid synthetic peptide derived from a conserved region of the SU glycoprotein of feline immunodeficiency virus (FIV), i.e., 225EGPTLGNWAREIWATLFKKA244, bound the surface of FIV-permissive cells and inhibited FIV infection of CrFK and lymphoid cells. In this paper, we report, by the use of N- and C-terminus deleted synthetic analogs and by glycine scanning experiments that the minimal sequence needed for the full antiviral activity of the peptide maps in correspondence of amino acids 229LGNWAREIWATL240 and that either tryptophans residues at sequence position 232 or 237 are essential for such activity. Circular dichroism (CD) studies indicate that in the presence of a hydrophobic environment the 225E-A244 peptide adopts a structure containing an amphipathic alpha-helical segment of approximately 7 residues, corresponding to 2 helical turns, likely in correspondence of the sequence 231(N)WAREIW(A)238. Such a helical segment of FIV SU glycoprotein may play a role in viral envelope fusion role with the host cell membrane, thus proving critical for cell infection.

Gastroesophageal reflux and bronchial asthma: prevalence and effect of cisapride therapy.

We evaluated the prevalence of gastroesophageal reflux in 36 children, 22 (61.2%) male and 14 (38.8%) female (median age, 75.5 months; range, 18-178), with noncontrolled asthma by means of prolonged (22-24 h) esophageal pH monitoring. None of the children had gastrointestinal symptoms suggesting gastroesophageal reflux. Atopy was seen in 21 of 36 (58.3%) patients. Pathological gastroesophageal reflux was present in 27 (75%) children. All patients were given cisapride (0.2 mg/kg q.i.d.) for 3 months. A clinical and pharmacological score was determined, and a second pH-metric study was made at the end of the follow-up period. The following pH-metric parameters were evaluated: the total percentage of time pH was < 4, the number of reflux episodes, the number of reflux episodes lasting > 5 min, the length of the longest single reflux episode, and the percentage of time the esophageal pH was < 4 during sleep. The study was completed in 11 of 27 children. The percentage of time that esophageal pH was < 4 improved in nine of 11 (81.8%) patients (p = 0.013). The percentage of time that esophageal pH was < 4 during sleep showed the most significant decrease (p = 0.002) after treatment. Improvement in both clinical and pharmacological scores was highly significant (p < 0.0001) in 19 of 27 patients, eight of whom did not want to repeat the pH study. We conclude therefore that gastroesophageal reflux is frequently associated with noncontrolled asthma and that medical therapy for reflux may improve the further course of respiratory disease.

[Gastroesophageal reflux and respiratory pathology]. / Reflusso gastroesofageo e patologia respiratoria.

The prevalence of gastroesophageal reflux (GER) in 86 children with respiratory disease (recurrent pneumonia, chronic cough, bronchial asthma) has been evaluated by mean of prolonged (22-24 hours) esophageal pH-monitoring. The following parameters were evaluated: the total percentage of time pH < 4 and the percent time the esophageal pH was < 4 while sleeping. None of the children had gastrointestinal symptoms suggesting GER and no neurological disorder was noted in any of the studied patients. The mean age was 68.98 +/- 46.46 months (range 14-189); 53 (61.6%) males and 33 (38.4%) females were considered in the study. Atopy was evidenced in 42/86 (48.8%) children (total IgE > 2SD in 42/86 and prick tests positiveness in 32/86. A pH-metry indicating pathological GER was present in 52/86 (60.5%) children: 39/62 (62.9%) patients with bronchial asthma, 5/10 (50%) subjects with chronic cough and 8/14 (57.2%) children with recurrent pneumonia. No significant difference in the diagnosis of GER was recorded between atopic or non-atopic patients. The children with abnormal pH-metric recording were also evaluated by upper gastrointestinal series and/or endoscopy. A conventional barium radiology was performed in 44/52 patients and confirmed GER in 19/44 (43.2%). Esophagitis was evidenced in 21/46 (45.7%) studied patients. The presence of esophagitis was significantly (p = 0.032) related to the total percentage of time pH < 4, but the most significant (p = 0.002) association was with the percent time the esophageal pH was < 4 during sleep.(ABSTRACT TRUNCATED AT 250 WORDS)

Five-year follow-up of vaccination against hepatitis B virus in newborns vaccinated with a reduced number of doses.

The protective efficacy and long-term immunogenicity of a schedule with a reduced number of vaccinations in infants born to mothers positive for hepatitis B surface antigen (HBsAg) has been investigated: 494 infants immunized with a plasma derived hepatitis B vaccine (Hevac B Pasteur) at day 20 after birth and 2 months later were studied. Anti-HBs seroconversion rate was 96% after the second dose, a value similar to that obtained in older children. At 5 years of life anti-HBs at protective levels was still 95.8%. In conclusion, we suggest that this effective and economical reduced vaccination schedule, which before this study was applied to older children, can be used in newborns from HBsAg positive mothers with good protective efficacy shifting the beginning of active immunization from birth to day 20.

False diagnosis of non-A/non-B hepatitis hiding two cases of cystic fibrosis.

We report two cases of children followed for many years with an original diagnosis of non-A/non-B hepatitis. One child developed serious cirrhosis with portal hypertension. Long-term observation of the course of their hepatic disease provided the diagnosis of cystic fibrosis. These cases demonstrate that cystic fibrosis though rarely presenting with initial hepatic signs, can manifest itself with only long-term hepatic symptoms. We therefore strongly recommend analysis of sweat chloride concentration in cases of hepatic disease of unknown origin.

Modulation by human milk of IgG subclass response to hepatitis B vaccine in infants.

The influence of breast and formula feeding on specific anti-hepatitis B surface antigen (HBsAg) IgG subclass production and distribution has been investigated in 40 healthy infants, born to HBsAg-positive mothers and vaccinated against hepatitis B virus (HBV). Twenty children were bottle fed and 20 were breast fed. Specific subclasses were detected at the 4th and 12th months using an enzyme-linked immunosorbent assay with monoclonal antibodies. A defect in total IgG and IgG subclasses was previously excluded. Significant differences were observed both at the 4th and 12th months for IgG1 and IgG2. Breast-fed infants had significantly higher levels of specific IgG2 (about three times higher), while IgG1 levels were significantly higher in formula-fed infants. Anti-HBsAg IgG4 levels were always higher in bottle-fed infants, but a statistical significance was never present. No difference was found in specific IgG3 levels. This study reports the evidence that breast feeding influences specific IgG subclass synthesis against a viral antigen and suggests an immunologic modulation of the response to vaccines dependent not only on age but also on factors present in human milk.

[Subclasses of IgG: biological aspects and functional behavior in response to vaccination]. / Le sottoclassi delle IgG: aspetti biologici e comportamento funzionale in risposta alle vaccinazioni.

The IgG subclasses are known to have different structure and functions. The IgG1 and IgG3 bind to monocyte and neutrophils and activate the complement more easily then IgG2 and IgG4. The levels of IgG subclasses found in newborns are mainly determined by the transplacental passage since the synthesis, in the first time of life, is very low. The levels found in adults are reached only during the adolescence. The immune response to a protein antigen is mainly in the IgG1 subclass, on the contrary the response to polysaccharide antigens is mainly IgG2. For that reason children, who produce very few IgG2 till they are 2 years old, cannot be vaccinated with carbohydrate vaccines unless a protein conjugate vaccine is used. In addition, the route of subministration, the dose of antigen, the age of vaccinated people and genetic factors can modify the subclass pattern obtained in response to vaccinations. For these reasons, probably, the immune response studied after a vaccination or in individuals who recovered from a natural disease is not always the same. Since the role of the different subclasses is not yet completely clarified, the interpretation of the importance of a selective "choice" of a subclass instead of another after vaccination is still difficult.

[Evaluation of the respiratory function in patients with Duchenne's muscular dystrophy]. / Valutazione della funzionalità respiratoria in pazienti affetti da distrofia muscolare di Duchenne.

The purpose of this investigation was to compare the respiratory function of patients with different clinical stages of Duchenne muscular dystrophy (DMD). Twenty-three DMD patients who were followed at Paediatric Department of Florence, were studied. We found a good correlation between the subject's functional capacity and normal predicted values for forced vital capacity (FVC) as well for forced volume at first second (FEV1). Subjects whose FVCs ranging from 10% to 20% were considered mildly involved; subjects with FVCs ranging from 20 to 30% were considered moderately involved, severe involved subjects with FVCs ranging from 30% to 40%. When the FVCs were less than 40% of the predicted values, the subjects were considered to have very severe respiratory impairment. In our study 83% of patients wheelchair bound was characterised by a very severe deterioration of pulmonary function.

[Robinow syndrome. Description of a case with diaphragmatic relaxation]. / La sindrome di Robinow. Descrizione di un caso con relaxatio diaframmatica.

The authors report a case of Robinow Syndrome in a child 22 months old, who presented a relaxatio diaphragmatic too. This is the first case, among about twenty cases indicated in literature, in whom this type of malformation is present.

[Infection by Mycoplasma pneumoniae. Epidemiologic investigation and statistical clinical study of 122 children admitted to the Institute of Pediatrics of the University of Florence during the period 1 January 1977 to 31 December 1983]. / L'infezione da Mycoplasma pneumoniae. Indagine epidemiologica e studio clinico statistico su 122 bambini ricoverati presso l'Istituto di Pediatria dell'Università di Firenze nel periodo 1 gennaio 1977-31 dicembre 1983.

We carried out an epidemiologic research about M. Pneumoniae infection from 1977 to 1983: determination of CF antibodies has been performed by the Bacteriology and Virology Laboratory (Prof. Lamanna, USL 10/D, Florence). Serum samples were collected by the laboratory itself and by hospitals and university departments of Florence and neighbouring communes. Our research confirms cyclic behaviour of M. Pneumoniae infection: we could follow up a 3 years long epidemic after a previous 2 years long endemic disease. We found a major sera-positive rate for M. Pneumoniae in children than in adults. In the same period we carried out a clinical-statistical study on 122 children (80 males and 42 females) admitted to the Pediatric Institute "A. Meyer" in Florence because of an infection due to M. Pneumoniae. The most affected age range sems to be included between 5 and 10 years. Pneumonia is the commonest clinical feature of M. Pneumoniae infection (85.2%): furthermore, we must point out some cases in 0-2 years-old subjects. In our patients, clinical features, X-ray findings and laboratory tests strictly agree with those reported by other authors. We treated all our patients with erythromycin: in three weeks we obtained thee normalization of clinical patterns and X-ray findings in all cases.

[Blood myoglobin in children with progressive muscular dystrophy and in carriers]. / La mioglobinemia in bambini affetti da distrofia muscolare progressiva e portatrici.

We studied the behaviour of serum myoglobin in 32 children affected by Duchenne muscular dystrophy, in 30 mothers (10 definite carriers and 20 possible carriers), in 5 sisters (possible carriers) and in 40 healthy women (control). The serum myoglobin was always increased in the patients affected by Duchenne muscular dystrophy; the greatest values were in the patients who were still ambulant, with a behaviour similar to creatine kinase. In the carriers the myoglobinemia showed a significant increase in definite carriers, while there was no significant difference between the possible carriers and the controls.

[Granulocyte function in Duchenne's progressive muscular dystrophy]. / La funzionalità granulocitaria nella distrofia muscolare progressiva di Duchenne.

We studied the in vitro function of polymorphonuclear leukocytes in 14 patients affected by Duchenne muscular dystrophy, aged 2-14 years, especially the chemotaxis and the bactericidal activity. The chemotaxis appeared significantly reduced, while the bactericidal activity was sometimes reduced, sometimes increased. In this disease the polymorphonuclear leukocytes seems to have a reduced capacity of movement; probably this phenomenon is connected with an alteration of the muscular cellular apparatus, which perhaps is related with an electrolytic intercellular inequality. These findings are consistent with the definition of Duchenne muscular dystrophy as a systemic disease of the membrane with alterations not only of striped muscular fibers, but of various cells and tissues.

[In-vitro block of granulocyte chemotaxis with attenuated measles virus]. / Blocco "in vitro" della chemiotassi granulocitaria mediato da virus attenuato del morbillo.

Live attenuated measles virus preparations, which are in use today in the prophylaxis against measles, were added in vitro to neutrophils from healthy children with negative history both for measles infection and vaccination. After incubation at 37 degree C for 30 min, and washing in Hank's solution, chemotaxis was assayed in Sykes-Moore chambers. Bacterial endotoxin and sera from healthy donors were used to stimulate motility. A significant decrease in chemotaxis was shown by virus-incubated neutrophils as compared to controls. These findings demonstrate that the attenuated measles virus preserves the same ability to inhibit neutrophil chemotaxis than the wild one, during the infection by which an in vivo defective chemotaxis has been previously described. It is suggested that some interference(s) of the virus may occur directly in the cell and/or its membrane receptors.

[Structural and ultrastructural features of the skeletal muscle in Werdnig-Hoffmann disease]. / Aspetti strutturali ed ultrastrutturali del muscolo striato nella malattia di Werdnig-Hoffmann.

In researches performed on striated muscles of individuals affected by various myopathies, fragments of gastrocnemius muscle in course of Werdnig-Hoffmann disease have been studied. The light microscopic observations demonstrate the presence of numerous, very thin, atrophic fibres with many irregularly lobated and packed nuclei alternating with apparently normal fibres with lighter and larger nuclei. The connective part of the muscle appears to be remarkably increased and replaces the atypical contractile tissue. The electron microscope shows, in the atrophic fibres, remarkable alterations of the sarcoplasmatic elements, and a complete disarrangement of myofibrils and myofilaments, some of which are anchored to fragments of the Z bands, irregularly arranged. Even the normal orientation of the sarcomeres often with indefinite bound, is upset. The numerous lobated nuclei totally occupy the irregular protrusions of the sarcoplasm, rich in peripheral microvesicles. In respect of other examined myopathies, in which the muscle fibres alterations affect limited parts of the contractile elements, in these cases the modifications seem to be of greater entity and to affect completely single muscle fibres.

[Immunological disorders in a patient with Whitaker syndrome (author's transl)]. / Alterazioni immunologiche in un caso di sindrome di Whitaker.

Some immunological parameters have been examinated in a 13-year-old girl with Whitaker syndrome and in her closest blood-relatives. Absolute and percentage values of T and B lyphocytes were normal, whereas mitogen responsiveness was over normal range before therapy had begun. This phenomen did not appear if the cultures were carried out in presence of autologous serum. Neutrophil function was defective in responsiveness to chemotactic stimulation, but NBT reduction as well as bactericidal activity were normal. Defects both in mitogen responsiveness and chemotaxis have been pointed out in the parents and in the patient's twin brother. The immunologic disorders observed are discussed in the light of the clinical and functional pecularities of Whitaker's syndrome.

[Xeroradiography in the investigation of muscular diseases in pediatric age (author's transl)]. / L'indagine xeroradiografica nello studio delle miopatie in età pediatrica.

A comparison is made between xeroradiography and traditional radiography in 43 patients suffering from myopathies in pediatric age. Only eight of the patients had a neurological genesis. Xeroradiography results to be much more useful in the early diagnosis of muscular disorders and in the correct definition of the disease.