Outcome of noninvasive ventilation in children with neuromuscular disease.

OBJECTIVE: To assess the effect of institution of noninvasive ventilation (NIV) on clinical outcome and quality of life (QOL) in a cohort of children with severe neuromuscular disorders. METHODS: We reviewed records and obtained clinical data from the year prior to commencing NIV and annually thereafter. Data obtained included diagnosis, patient symptoms, mortality, NIV adverse effects, pulmonary function tests, polysomnographic data, length of hospitalizations, and health care costs. Patients and parents completed questionnaires assessing QOL with NIV and recalling QOL before NIV. RESULTS: Fourteen of 17 (82%) suitable patients were enrolled. Follow-up ranged from 6 to 84 months (median 30). Symptoms of daytime sleepiness (p = 0.003) and headache (p = 0.046) improved after initiation of NIV. Sleep quality assessed by polysomnography also improved. Hospitalization rates (p = 0.002) and health care costs (p = 0.003) decreased. QOL remained stable after NIV, despite disease progression. CONCLUSION: Treatment of respiratory failure, in children with neuromuscular disease, with noninvasive ventilation results in a reduction in symptoms, hospitalizations, and health care costs without adverse effects on quality of life.

Characterisation of OXA-51, a novel class D carbapenemase found in genetically unrelated clinical strains of Acinetobacter baumannii from Argentina.

Acinetobacter baumannii is now one of the most frequently encountered nosocomial pathogens in intensive therapy units, and is renowned for being difficult to treat because of resistance to most antibiotics. Carbapenems are the remaining drugs of choice in many centres, but carbapenem resistance is now emerging in strains worldwide. Two subgroups of carbapenem-hydrolysing beta-lactamases, which differ in their amino-acid homology, have been found in some resistant strains. This report describes the emergence and characterisation of a novel carbapenemase (OXA-51) in genetically distinct carbapenem-resistant A. baumannii strains from Argentina. Enzyme kinetics and inhibitor studies were performed spectrophotometrically with purified beta-lactamase. Amplification of the gene was achieved with a two-step PCR method employing arbitrary partially degenerate and gene-specific primers. Transfer of imipenem resistance was attempted with the use of broth and membrane filter methods. Attempts to produce plasmid-cured variants were made in ethidium bromide curing experiments. OXA-51 was identified in two clones of A. baumannii, and was found to have < 63% amino-acid identity with subgroups 1 and 2. Enzyme kinetic studies confirmed that OXA-51 was a molecular class D enzyme with carbapenemase activity, and that it displayed the highest affinity for imipenem (Km value 11 microM). Sequence analysis of the gene identified distinct differences within conserved class D motifs when compared with subgroups 1 and 2. Attempts to transfer imipenem resistance and to determine a plasmid location for the gene failed. OXA-51 is the first of a new subgroup of carbapenemases to emerge in multiresistant clinical isolates of A. baumannii.

[Presence of integrons and their relationships with the resistance to third generation cephalosporins among nosocomial isolates of Acinetobacter baumannii]. / Presencia de integrones y su relación con la resistencia a cefalosporinas de tercera generación en cepas de Acinetobacter baumannii de origen nosocomial.

BACKGROUND: Acinetobacter baumannii is an important etiological agent causing nosocomial infections. High level of resistance for different kind of antimicrobials has been observed, including beta-lactam antibiotics. This feature, chromosomal or plasmid encoded, has been associated to integrons harbouring antibiotic resistance gene cassettes. AIMS: To investigate the presence of integrons among clinical isolates resistant to third generation cephalosporins (3GC). MATERIAL AND METHODS: One hundred A. baumannii strains isolated from several Chilean hospitals were included in this study. Minimal inhibitory concentrations (MIC) of 3GC by an agar dilution method were carried out. Integrons class 1, 2 and 3 were investigated by colony blot hybridisation and confirmed by PCR. RESULTS: High level of resistance to all assayed 3GC was observed. On the other hand, integrón class 2 was the most prevalent (77% of isolates) followed by integron class 1 (52%). Forty six percent of isolates hybridised with probes for both of them. However, no positive hybridisation was detected for integron class 3. CONCLUSIONS: Nevertheless, most isolates harboured one or both class of integron; there was no direct relationship between the presence of these genetic structures and the resistance to this kind of beta-lactam antibiotics.

Identification and characterization of vanB2 glycopeptide resistance elements in enterococci isolated in Scotland.

Thirty-two vanB glycopeptide-resistant enterococci (28 Enterococcus faecium and 4 Enterococcus faecalis) were collected from hospitalized patients in Glasgow, Edinburgh, Dundee, and Aberdeen, Scotland, and the vanB element in each was compared to vanB1 of E. faecalis strain ATCC 51299. HhaI digestion of PCR fragments of the vanB ligase gene was used to identify vanB subtypes. All E. faecium isolates were vanB2, and all E. faecalis isolates were vanB1. Restriction fragment length polymorphism analysis of a 5,180-bp vanS(B)-vanX(B) long-PCR fragment of the vanB cluster showed the loss of HaeII restriction sites in vanS(B), vanW, and vanX(B) in strains containing a vanB2 ligase gene. Partial sequences of genes in the vanB2 cluster for two genomically distinct Scottish isolates were >99.8% identical to each other. vanS(B2), vanX(B2), and vanB2 sequences differed at the nucleotide level from those of vanS(B), vanX(B), and vanB by 4.2, 4.6, and 4.8%, respectively. The vanB2 resistance element appears to be widespread among VanB glycopeptide-resistant E. faecium strains isolated in Scottish hospitals.

Sequence analysis of ARI-1, a novel OXA beta-lactamase, responsible for imipenem resistance in Acinetobacter baumannii 6B92.

The sequence of the bla(ARI-1) gene from imipenem-resistant Acinetobacter baumannii 6B92 has been determined. The structural gene encodes a 273-amino-acid protein which is most related to the OXA class D beta-lactamases. The conserved S-T-F-K and K-T-G motifs were identified in the ARI-1 protein sequence, also named OXA-23, but significantly, a point mutation (Y-->F) was identified in the Y-G-N conserved motif, also known to function in the active site.

Identification and characterization of class 1 integrons in bacteria from an aquatic environment.

In a survey of 3000 Gram-negative bacteria isolated from an estuarine environment over a 2 month period, the incidence of class 1 integrons was determined to be 3.6%. Of 85 integrons studied further, 11 lacked both the qacEdelta1 and sull genes usually present in the 3' conserved segment of the integron. The qacEdelta1 and sull genes were identified in the 3' conserved segment of 36 integrons. The remaining 38 integrons lacked a sull gene but contained a qacE gene. The variable region of 74 integrons was characterized by PCR and sequence analysis. Forty of the integrons were found to lack integrated gene cassettes, although 21 of these 'empty' integrons were shown to contain inserted DNA which has been tentatively identified as a novel insertion sequence (IS) element. Of the 34 integrons which contained inserted gene cassettes, the aadA1a gene was found to be the most prevalent (74%). Nineteen integrons contained additional or other gene cassettes in their variable region, including those encoding resistance to trimethoprim (dfr1a, dfrIIc, dfrV, dfrVII, dfrXII), chloramphenicol (catB3, catB5), aminoglycosides (aadA2, aacA4, aacC1), beta-lactamases (oxa2) and erythromycin (ereA). This study confirms the occurrence of integrons in bacteria from a natural habitat and suggests that in the absence of continued antibiotic selective pressures, integrons which persist appear to preferentially exist without integrated antibiotic resistance gene cassettes.

Evaluation of nitrone spin-trapping agents as radioprotectors.

The focus of this investigation was to determine whether the nitrone spin-trapping compounds alpha-phenyl-N-tert-butylnitrone (PBN), 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) and alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone (POBN) are radioprotectors. Two methods were used to assess for radioprotection: measurement of oxidative damage to DNA bases and mammalian cell survival assays. Oxidative damage to DNA was quantified by measuring the relative amounts of 8-hydroxy-2'-deoxyguanosine (8-OH-dG) produced by the reaction of hydroxyl radicals (OH.) with 2-deoxyguanosine (dG) after irradiation. PBN, DMPO and POBN, when dissolved in aqueous solutions of either dG or naked salmon sperm DNA, reduced the formation of 8-OH-dG by 137Cs gamma irradiation significantly. The spin-trapping agents, especially PBN at lower concentrations, were more effective in preventing radiation-induced formation of 8-OH-dG in naked DNA than in free dG. These data suggest that PBN, DMPO and POBN act as free radical scavengers which may associate with DNA and afford protection against gamma rays. However, no enhancement of survival was observed when Chinese hamster ovary (CHO) cells were exposed to high non-toxic concentrations of PBN or POBN prior to and during irradiation with 60Co gamma rays and scored for clonogenic survival. DMPO provided only minimal protection from radiation-induced cell killing.

Nucleotide sequence and genetic analysis of the type Ib trimethoprim-resistant, Tn4132-encoded dihydrofolate reductase.

Transposon Tn4132, encoding the type Ib trimethoprim-resistant, dihydrofolate reductase, was transposed from the clinical plasmid pUK163 to a small recombinant plasmid pZMR12. Restriction endonuclease and partial sequence analysis of Tn4132 revealed a close relationship to Tn7. The nucleotide sequence of the dhfrIb trimethoprim-resistance gene was determined and the gene and its product were found to share significant homology with the dhfrIa, V, VI and VII plasmid-encoded dihydrofolate reductases. Extensive sequence homology (88%) was observed with the type V dihydrofolate reductase at both the nucleotide and amino acid level. Oligonucleotide probes, distinguishing between the dhfrIb and dhfrV genes, were designed. The discriminatory capabilities of these probes in future epidemiological studies will permit a more accurate determination of the dissemination of these two closely related trimethoprim-resistance genes than has previously been possible.

Trimethoprim resistance in urinary pathogens in northern Scotland: epidemic spread of a resistance plasmid encoding the type Ib trimethoprim-resistant dihydrofolate reductase.

The prevalence of trimethoprim resistance in enterobacterial urinary pathogens from hospitalised patients in the Angus district of northern Scotland (22.8%) was twice that found in similar isolates from patients attending general practitioners (11.2%). Thirty-three of the 143 trimethoprim-resistant strains were shown to harbour transferable plasmids conferring high-level trimethoprim resistance. In total, 17 different plasmid types were distinguished. Two plasmids, pUK1184 and pUK1185, accounted for 36% of the trimethoprim resistance plasmids and were shown by restriction endonuclease digestion fingerprints to be closely related to plasmid pUK28, previously demonstrated to be endemic in urinary pathogens in the Edinburgh area. Only 21% of the plasmids were shown to encode the type Ia trimethoprim-resistant dihydrofolate reductase, whereas 70% of the trimethoprim resistance plasmids were found to encode the type Ib dihydrofolate reductase. Hybridisation of the trimethoprim resistance plasmids identified in this study with gene probes specific for the integrase genes of transposons Tn7 and Tn21 indicates that the dhfrIa is rarely present within Tn7 or related transposons in these plasmids and may be more prevalent within Tn21-like transposons. In contrast, with the exception of the two endemic plasmids that harboured the dhfrIb gene within a Tn7-like transposon, the majority of dhfrIb genes were not found to be associated with either Tn7- or Tn21-like structures.

Do nonclinical uses of antibiotics make a difference?

An increasing range of antibacterial compounds is being used for nonclinical purposes, especially in the fields of animal husbandry and fish farming. As in human medicine, exposure to antibiotics has lead to the emergence of antibiotic-resistant bacteria in animal populations. The potential impact of antibiotic use in animals on human health and the management of clinical infections in humans is discussed in light of growing evidence to suggest that "new" resistance genes and multiresistant pathogens with increased pathogenicity are emerging in food animals as a direct consequence of antibiotic exposure.

Antimicrobial resistance spread in aquatic environments.

The increased use of antimicrobials in farming, together with the practice of raw sewage discharge into receiving waters, has resulted in a significant increase in the numbers of antibiotic resistant bacteria present in aquatic environments. The role of this environment to act, not only as a reservoir of clinical resistance genes, but also as a medium for the spread and evolution of resistance genes and their vectors, is discussed.

The role of thymine starvation in the expression of type IV plasmid-encoded trimethoprim-resistant dihydrofolate reductase.

Hyperproduction of the type IV plasmid-encoded dihydrofolate reductase was studied in Escherichia coli J62-2 (pUK1123). Hyperproduction of the enzyme was shown to occur not simply as a response to a given concentration of trimethoprim but also to the presence of thymidine in the medium. Before hyperproduction occurred the bacteria began to elongate and die, thus showing the symptoms of thymine starvation. Hyperproduction also required the presence of L-methionine, adenine and glycine, suggesting that the elevated production of the enzyme was a response to the ability of trimethoprim to starve the cell of thymine metabolites.

The induction of trimethoprim resistance encoded by the type IV dihydrofolate reductase gene.

The effect of plasmid pUK1123, which confers low level resistance to trimethoprim when tested on solid minimal medium, but also no resistance when tested on IsoSensitest agar, was investigated in liquid media. The growth of Escherichia coli J62-2, harbouring pUK1123, was unaffected in liquid minimal medium containing trimethoprim 10 mg/L. However, in IsoSensitest broth, exposure to this drug concentration resulted in bacteriostasis. After an initial delay, resistance to trimethoprim was induced in IsoSensitest broth containing trimethoprim 10 mg/L, by the imposition of thymine starvation. This response was immediately reversible when trimethoprim was removed, confirming that resistance resulted from induction rather than selection of resistant mutants.

The incidence of antibiotic resistance in aerobic faecal flora in south India.

During a field study in South India in 1989, faecal specimens were collected from residents in villages and the town of Vellore in South India. Examination of the faecal specimens revealed that virtually the whole population carried commensal bacteria resistant to trimethoprim, ampicillin and chloramphenicol. Most specimens contained more than one type of bacterium resistant to each antibiotic. There was less resistance to nalidixic acid, with a higher proportion in the town (33%) than in the villages (13%). Although there was little cross-resistance of the ampicillin-resistant strains to later generation cephalosporins, 50% were resistant to the combination of amoxycillin and clavulanic acid. There was no significant cross-resistance of the nalidixic acid-resistant strains to fluorinated 4-quinolones, despite the free availability of ciprofloxacin and norfloxacin in the area. The probable reason for the high incidence of resistance to first generation antimicrobials is the extensive use of these agents, coupled with continuous exposure to large numbers of faecal micro-organisms.