Safety of ultrashort-term sit with pollen allergoids adjuvanted by monophosphoryl lipid A: a prospective Italian survey.

A 3-year prospective post marketing survey on the safety of the recently developed ultrashort pre-seasonal subcutaneous immunotherapy (uSCIT-MPL4) with pollen allergoids adjuvanted with monophosphoryl lipid A was performed. A total of 510 patients received uSCIT-MPL4, 61% for grass, 35.7% for birch, 13.2% for parietaria and 3% for other pollens (ragweed, mugwort, and olive). A total of 3308 injections were given and the mean duration of uSCIT-MPL-4 was 2.3 years. Overall, only 7 slight systemic reactions (SR) were observed in 510 patients (1.37%) and 2.11/1000 injections suggesting that this treatment is even safer than traditional depot injection SIT.

Pharmacogenetics and pharmacogenomics: are they still promising?

In the last several years pharmacogenetics and pharmacogenomics have attracted the interest of the scientific community and of important pharmaceutical groups. What is the consequence for medicine and for the pharmaceutical industry? What has emerged from this investment, and what can we expect for the future? As with many new technologies, pharmacogenetics and pharmacogenomics were first adapted with much enthusiasm, and then found to require time and experience, together with sustained investment, before they could take their full place in drug discovery and development. The benefits of these technologies are now emerging, however, and they have become essential tools for the pharmaceutical industry.

Family of class 1 integrons related to In4 from Tn1696.

The class 1 integron In28, found in the multidrug resistance transposon Tn1403, was found to be located in the res site of the backbone transposon and is flanked by a 5-bp direct duplication, indicating that it reached this position by transposition. In28 has a backbone structure related to that of In4, but has lost internal sequences, including the sul1 gene, due to an IS6100-mediated deletion. In28 also lacks the partial copy of IS6100 found in In4 and contains different gene cassettes, blaP1, cmlA1, and aadA1. In1, the class 1 integron found in the multidrug resistance plasmid R46, is also located in a putative res site and belongs to the In4 group. In1 has a shorter internal deletion than In28 and has also lost one end. Additional integrons with structures related to In4 were also found in databases, and most of them had also lost either one end or internal regions or both. Tn610 belongs to this group.

Diaminodiphenylmethane (DDM): frequency of sensitization, clinical relevance and concomitant positive reactions.

Diaminodiphenylmethane (DDM) is an aromatic diamine used in the manufacture of rubber, plastics, diisocyanates, dyes and adhesives. It may cross-react with para-(amino)compounds. Allergic patch test reactions to DDM are relatively frequent, but their relevance is often difficult to detect. We report our experience in 6809 patients (4589 female, 2220 male, mean age 39.9+/-17.8 years) with suspected contact dermatitis patch tested during the period 1997-1999 by the North-East Italy Contact Dermatitis Group (NEICDG). A positive patch test to DDM was detected in 132 (1.9%) patients (88 female, 44 male, mean age 49.5+/-16.2 years). Eczema was mostly localized on the hands. The relevance was detected in 31 patients. A logistic regression analysis showed an association with patient's age (odds ratio 5.4 for age 30-59 years), absence of atopic diseases (odds ratio 3.1) and presence of leg ulcer (odds ratio 5). We found a highly significant correlation (p<0.001) between sensitivity to DDM and to para-phenylenediamine, Disperse Yellow 3, cobalt chloride, fragrance mix, benzocaine, paraben mix and primin. Positive patch test results to DDM were relatively frequent. The difficulty in detecting the relevance of these sensitizations may be related to the surprisingly high frequency of concomitant positive reactions to other allergens.

Efficiency of recombination reactions catalyzed by class 1 integron integrase IntI1.

The class 1 integron integrase, IntI1, recognizes two distinct types of recombination sites, attI sites, found in integrons, and members of the 59-be family, found in gene cassettes. The efficiencies of the integrative version of the three possible reactions, i.e., between two 59-be, between attI1 and a 59-be, or between two attI1 sites, were compared. Recombination events involving two attI1 sites were significantly less efficient than the reactions in which a 59-be participated, and the attI1 x 59-be reaction was generally preferred over the 59-be x 59-be reaction. Recombination of attI1 with secondary sites was less efficient than the 59-be x secondary site reaction.

Recovery of new integron classes from environmental DNA.

Integrons are genetic elements known for their role in the acquisition and expression of genes conferring antibiotic resistance. Such acquisition is mediated by an integron-encoded integrase, which captures genes that are part of gene cassettes. To test whether integrons occur in environments with no known history of antibiotic exposure, PCR primers were designed to conserved regions of the integrase gene and the gene cassette recombination site. Amplicons generated from four environmental DNA samples contained features typical of the integrons found in antibiotic-resistant and pathogenic bacteria. The sequence diversity of the integrase genes in these clones was sufficient to classify them within three new classes of integron. Since they are derived from environments not associated with antibiotic use, integrons appear to be more prevalent in bacteria than previously observed.

Resolution of holliday junctions by RuvABC prevents dimer formation in rep mutants and UV-irradiated cells.

In this work, we present evidence that indicates that RuvABC proteins resolve Holliday junctions in a way that prevents dimer formation in vivo. First, although arrested replication forks are rescued by recombinational repair in cells deficient for the Rep helicase, rep mutants do not require the XerCD proteins or the dif site for viability. This shows that the recombination events at arrested replication forks are generally not accompanied by the formation of chromosome dimers. Secondly, resolution of dimers into monomers is essential in the rep ruv strain because of an increased frequency of RecFOR recombination events in the chromosome of this mutant. This suggests that, in the absence of the Ruv proteins, chromosomal recombination leads to frequent dimerization. Thirdly, dif or xerC mutations increase the UV sensitivity of ruv-deficient cells 100-fold, whereas they do not confer UV sensitivity to ruv+ cells. This shows that recombinational repair of UV lesions is not accompanied by dimer formation provided that the RuvABC proteins are active. The requirement for dimer resolution in ruv strains is suppressed by the expression of the RusA Holliday junction resolvase; therefore, RusA also prevents dimer formation. We conclude that the inviability arising from a high frequency of dimer formation in rep or UV-irradiated cells is only observed in the absence of known enzymes that resolve Holliday junctions.

An epidemiological study to assess migraine prevalence in a sample of Italian population presenting to their GPs.

This multicentre, observational, cross-sectional study was conducted to determine migraine prevalence in a sample of population presenting to their GPs. The study covered all the patients who visited the GPs practice, for any reason, on 5 consecutive days of 2 different weeks. A total of 71,588 patients were interviewed by 902 GPs. The prevalence of migraine in this sample was 11.6%.

Rise and fall of asthma-related mortality in Italy and sales of beta2-agonists, 1980-1994.

We performed this study with the aims of describing the trend of asthma-related mortality in Italy between 1980 and 1994, and to evaluate the relationship between sale estimates of beta2-agonists drugs and mortality from asthma. For asthma mortality we used data provided by National Institute of Statistics, for sale estimates of beta2-agonists we used data provided by IMS HEALTH. We calculated the gender specific age-standardized incidence rates of asthma-related deaths for all ages and for age classes. We found that estimates for asthma-related mortality steadily increased between 1980 and 1987 in both sexes, and thereafter decreased. In people, aged between 34 and 64 and over 64, death rates in males were significantly higher than in females while the rates in those aged less than 34, were mostly similar in both gender. The overall exposure to beta2-agonists (alone and in combination) increased from 1980 to 1990, remained stable between 1990 and 1993, and increased steeply in 1994. We conclude that asthma-related death rates have declined since the mid-1980's. This decline has been more pronounced in males and in the older ages, while the rates in younger patients of both genders have remained nearly unchanged. Our data do not substantiate the hypothesis of an increased risk of asthma-related mortality associated to the use of inhaled beta2-agonists in general nor fenoterol or salbutamol in particular.

FtsK-dependent and -independent pathways of Xer site-specific recombination.

Homologous recombination between circular chromosomes generates dimers that cannot be segregated at cell division. Escherichia coli Xer site-specific recombination converts chromosomal and plasmid dimers to monomers. Two recombinases, XerC and XerD, act at the E. coli chromosomal recombination site, dif, and at related sites in plasmids. We demonstrate that Xer recombination at plasmid dif sites occurs efficiently only when FtsK is present and under conditions that allow chromosomal dimer formation, whereas recombination at the plasmid sites cer and psi is independent of these factors. We propose that the chromosome dimer- and FtsK-dependent process that activates Xer recombination at plasmid dif also activates Xer recombination at chromosomal dif. The defects in chromosome segregation that result from mutation of the FtsK C-terminus are attributable to the failure of Xer recombination to resolve chromosome dimers to monomers. Conditions that lead to FtsK-independent Xer recombination support the hypothesis that FtsK acts on Holliday junction Xer recombination intermediates.

Mobile gene cassettes and integrons in evolution.

Integrons and the site-specific recombination systems encoded by them provide a simple mechanism for the addition of new genes to bacterial chromosomes. Although there is substantial divergence among the four known integron-encoded integrases, they all recognize the recombination sites, known as 59-base elements, that are associated with genes that are packaged in gene cassettes. In contrast, the integron-associated recombination sites, attl sites, are preferentially recognized by the cognate integrase.

Measuring quality of life in dyspeptic patients: development and validation of a new specific health status questionnaire: final report from the Italian QPD project involving 4000 patients.

OBJECTIVE: Despite the fact that gastrointestinal disorders represent one of the most common reasons for medical consultations, formal assessment of patients' health-related quality of life (HRQOL) has been carried out only in a few studies, and in most cases generic questionnaires have been adopted. Because the specific issue of living with dyspeptic problems has been addressed in very few cases and no questionnaire has been shown to be appropriate for the Italian setting, a prospective project was launched to develop a specific HRQOL questionnaire for dyspepsia sufferers tailored to Italian patients but also appropriate in other cultural settings. METHODS: The project consisted in a 3-yr, three-phase survey, in which different versions of the quality of life in peptic disease questionnaire (QPD) were developed through expert and patient focus groups and empiric field studies and then administered to patients recruited in five multicenter studies. Standard psychometric techniques were used to evaluate the validity, reliability, responsiveness, and patient acceptability of the QPD. RESULTS: Three different versions of the QPD questionnaire were self-administered to more than 4000 patients. The final 30-item version, measuring three health concepts related to dyspeptic disease (anxiety induced by pain, social restriction, symptom perception), fulfilled the recommended psychometric criteria in terms of reliability and validity, correlated with health concepts measured with a well-known independent generic HRQOL instrument (the SF-36 Health Survey questionnaire) and was relatively invariant to diagnosis and sociodemographic variables; it also correlated with a measure of gastric pain frequency and was able to detect meaningful differences over time. CONCLUSIONS: Although further validation studies in different cultural and linguistic settings are mandatory before any firm conclusions can be drawn regarding the cross-cultural validity of the QPD, the data obtained provide evidence of the psychometric validity and robustness of the questionnaire when used in a fairly large, well-characterized population of Italian dyspeptic patients.

[Outcome research: rationales and objectives]. / La ricerca di esito: razionali ed obiettivi.

Outcome research measures the clinical, economic and humanistic outcomes of pharmacological therapies on patients suffering from different diseases. Outcome research can be applied either to phase II and III clinical trials to assess new drugs for registration or to already marketed drugs to assess their real value when used in clinical practice in a large population of patients. Oncology can be a major field of application because there is a large amount of new treatments put on the market, often without demonstration of significant improvement in survival or Quality of Life but with higher prices than the old molecules. Patients and disease specific outcomes have been identified for cancer by a Working Group set up by the American Society of Clinical Oncology, with the former being defined as more critical to establish the value of new treatments. Survival, drug toxicity and impact on Quality of Life are the most important patient's outcomes to consider before recommending anti-cancer therapies for use in clinical practice. The present paper will describe the outcomes in oncology and will deal with the transferability to clinical practice of the results of clinical trials.

The epidemiology of migraine: a retrospective study in Italian emergency departments.

This study was conducted to measure the frequency of contact with emergency departments in Italy because of migraine, and to compare the initial diagnosi s of headache with the diagnosis after application of the International Headache Society (IHS) criteria. A retrospective observational method was used, consisting of an analysis of the records of patients admitted to nine Italian emergency departments during different 4-month periods in 1994. Comparison of the initial diagnosis with the diagnosis after application of the IHS diagnostic criteria was performed. More than 31 million emergency department contacts were reported in Italy during 1994. In the same year, 543 630 patients visited the nine emergency departments enrolled in the study, with 169 569 of these contacts occurring in the 4-month period analyzed in the study. We excluded from the analysis all cases of secondary headache fully recognized at the emergency department admission (ie, traumas, intracranial pathology, systemic diseases). The total number of patients included in our analysis was 1043 (0.6%). The 934 patients who could be fully evaluated were initially classified as having migraine; cluster headache; headache not otherwise specified; or diagnosed in the emergency department as suffering from headache, but reclassified by other departments as suffering from a different disease. After retrospective application of the IHS classification, the diagnostic distribution was modified, revealing that 18% of patients with migraine and 5% with cluster headaches had previously been classified as having headache not otherwise specified; a further 6% of cases with migraine and 0.4% of patients with cluster headache had previously been classified as having secondary headaches. The diagnosis of headache not otherwise specified was made with notable frequency, indicating the limits of emergency department logs and the difficulty in carrying out a retrospective analysis and reassessment of diagnosis. The majority (88%) of patients assessed had not taken drugs for headache in the 48 hours before the emergency department contact, suggesting that in Italy emergency departments are used instead of a visit to the general practitioner. Nonsteroidal anti-inflammatory drugs were the most frequently prescribed drugs in the emergency departments for this group of diagnoses. The research revealed, on the one hand, that headache is a numerically significant phenomenon in the emergency department setting and, on the other, the need to apply prospective designs to this kind of survey.

Origins of the mobile gene cassettes found in integrons.

Many of the acquired antibiotic resistance genes found in enterobacteria and pseudomonads are part of small mobile elements known as gene cassettes, and other genes are also likely to be found in cassettes. The origins of the genes and the recombination sites that make up cassettes are not known, but recent analyses of available data suggest that cassettes may be ancient structures, and some hypotheses for how they are formed can now be examined.

Structure and function of 59-base element recombination sites associated with mobile gene cassettes.

The integration of gene cassettes into integrons is effected by site-specific recombination catalysed by an integrase, IntI, encoded by the integron. The cassette-associated recombination sites, 59-base elements, are not highly conserved and vary in length from 57 to 141 bp. They can be identified by their location and the relationship of over 20 bp at their outer ends to consensus sequences that are imperfect inverted repeats of one another. The recombination cross-over occurs close to one end of the 59-base element, within a conserved core site with the consensus sequence GTTAGGC or GTTRRRY. By introducing single-base changes at each of these positions in the aadB 59-base element, bases that are critical for site activity were identified. The recombination cross-over was also localized to a unique position between the adjacent G and T residues. Changes introduced in the conserved AAC of the inverse core site (GCCTAAC or RYYYAAC) located at the opposite end of the 59-base element also reduced site activity but to a lesser extent. Sequences of rare recombinants revealed an alternative position for strand exchange and led to the conclusion that 59-base elements comprise two simple sites, analogous to those recognized by other integrases, with each simple site made up of a pair of inversely oriented IntI binding domains separated by a spacer of 7 or 8 bp. Re-examination of the sequences of all known 59-base elements revealed that this simple site configuration was present at both the left and right ends in all 59-base elements. The identity of bases in the spacer is not required for efficient recombination and the cross-over is located at one end of the spacer, suggesting that during IntI1-mediated recombination only one strand exchange occurs.

[Development and validation of QPD 32, a specific questionnaire for measuring the quality of life of patients with peptic ulcer]. / Sviluppo e validazione di un questionario specifico per la misurazione della qualità della vita nei pazienti con malattia peptica: QPD 32.

Drugs need to be evaluated both in terms of efficacy, safety and regarding the patient's perception of his own health status. For these reasons, sensible, reliable and patient-oriented instruments are needed, besides the methodologies for evaluation of drug efficacy and safety. Such instruments substantially evaluate Health related Quality of Life (HrQoL). Concerning gastric acid hypersecretion few papers are available, based on HrQoL questionnaires, both general and specific. A research project led us to develop through patients and physicians involvement, a specific instrument to evaluate HrQoL as to the various aspects of the peptic disease. The project started in 1993 through a series of 4 focus groups with gastroenterologists and patients, followed by the preparation of a questionnaire named QPD48. Such instrument was psychometrically validated through a study named Herqules 1, involving 176 gastroenterologists and 1774 patients. The psychometric analysis on QPD48 led to the re-issue of a questionnaire named QPD32 with Chronbach's alfa equal to 0.91, based on 3 factor-referenced subscales evaluating pain, induced anxiety, constrained daily living and awareness of symptoms and agents. Concerning the concurrent validity a one-way analysis of variance showed highly significant differences associated with attack frequency with substantial effect sizes ranging from 0.46 to 1.27 of a standard deviation in the full scale. QPD 32 is patent protected and will be used in clinical trials.

Plasmid evolution by acquisition of mobile gene cassettes: plasmid pIE723 contains the aadB gene cassette precisely inserted at a secondary site in the incQ plasmid RSF1010.

Gene cassettes are mobile DNA elements which contain a specific recombination site, a 59-base element, recognized by the site-specific recombination system of integrons. Gene cassettes are normally found inserted at a unique site in an integron, downstream of a promoter which directs transcription of the cassette-associated genes. However, insertion of a gene cassette into a secondary site in a plasmid which does not contain an integron is also formally possible. Sequence analysis of the aadB gene in pIE723, a plasmid closely related to the IncQ plasmid RSF1010, revealed the presence of the complete aadB cassette inserted at a secondary site downstream of a known RSF1010 promoter. The site of insertion of the aadB cassette in RSF1010 conformed to the consensus for secondary sites recognized by the integron integrase (Int), and it is likely that the cassette was inserted via a single Int-mediated recombination event between the 59-base element of a free, circular aadB cassette and a secondary site in RSF1010. The cassette-associated recombination site was inactivated by the insertion, and Int-mediated excision of the aadB cassette from this non-specific location was not detectable, indicating that the cassette is stably inserted. The movement of gene cassettes to secondary sites is likely to play an important role in the acquisition of new genes by bacterial and plasmid genomes.