Co-occurrence of the cephalosporinase cepA and carbapenemase cfiA genes in a Bacteroides fragilis division II strain, an unexpected finding.

BACKGROUND: Bacteroides fragilis, an anaerobic gut bacterium and opportunistic pathogen, comprises two genetically divergent groups (or divisions) at the species level. Differences exist both in the core and accessory genomes and the beta-lactamase genes, with the cephalosporinase gene cepA represented only in division I and the carbapenemase gene cfiA only in division II. METHODS: Multidrug resistance in a clinical B. fragilis strain was examined by whole-genome sequencing. RESULTS: Strain CNM20200260 carried the antimicrobial resistance genes cepA, cfiA2, ant(6'), erm(F), mef(En2), est(T), tet(Q) and cat(A), along with 82-Phe mutation in gyrA (together with 47 amino acid changes in gyrA/B and parC/parE). bexA/B and other efflux pump genes were also observed. None of the detected insertion sequences was located upstream of cfiA2. The genome-based taxonomy coefficients (average nucleotide identity, DNA-DNA hybridization similarity and difference in genomic G + C%) with respect to genomes of the strains of B. fragilis division II and the novel species Bacteroides hominis (both cfiA-positive) met the criteria for CNM20200260 to belong to either species (>95%, >70% and <1%, respectively). No such similarity was seen with type strain NCTC 9343 or the representative genome FDAARGOS 1225 of B. fragilis (division I, cfiA-negative). Strain CNM20200260 harboured four out of nine Kyoto Encyclopedia of Genes and Genomes orthologues defined for division I and one of two defined for division II. CONCLUSIONS: This is the first description of the co-occurrence of cepA and cfiA in a Bacteroides strain, confirming the complexity of the taxonomy of this species.

Evaluation of CHROMagar™ B. cepacia agar for the detection of Burkholderia cepacia complex species from sputum samples of patients with cystic fibrosis.

INTRODUCTION: Burkholderia cepacia complex (BCC) are non-fermenting Gram-negative bacteria that can chronically colonize the lungs of people with cystic fibrosis (pwCF), causing a severe and progressive respiratory failure, post-transplant complications and epidemic outbreaks. Therefore, rapid and accurate identification of these bacteria is relevant for pwCF, in order to facilitate early eradication and prevent chronic colonization. However, BCCs are often quite difficult to detect on culture media as they have a slow growth rate and can be hidden by other fast-growing microorganisms, including Pseudomonas aeruginosa and filamentous fungi. MATERIAL AND METHODS: We evaluated the sensitivity of CHROMagar™ B. cepacia agar using 11 isolates from a well-characterized BCC collection, using BCA agar (Oxoid, UK) as a gold standard. We also studied 180 clinical sputum samples to calculate positive (PPV) and negative (NPV) predictive values. Furthermore, we used three of the well-characterized BCC isolates to determine the limit of detection (LOD). RESULTS: Eleven isolates grew on CHROMagar™ B. cepacia at 37ºC after 48 h. The NPV and PPV of CHROMagar™ B. cepacia were 100% and 87.5%, respectively. The LOD of CHROMagar™ B. cepacia was around 1 × 103 CFU/ml, requiring a ten-fold dilution lower bacterial load than BCA for BCC detection. CONCLUSION: CHROMagar™ B. cepacia agar proved to have a very good sensitivity and specificity for the detection of clinical BCCs. Moreover, the chromogenic nature of the medium allowed us to clearly differentiate BCC from other Gram-negative species, filamentous fungi and yeasts, thereby facilitating the identification of contaminants.

Focusing on Gordonia Infections: Distribution, Antimicrobial Susceptibilities and Phylogeny.

The immunosuppression conditions and the presence of medical devices in patients favor the Gordonia infections. However, the features of this aerobic actinomycete have been little explored. Strains (n = 164) were characterized with 16S rDNA and secA1 genes to define their phylogenetic relationships, and subjected to broth microdilution to profile the antimicrobial susceptibilities of Gordonia species that caused infections in Spain during the 2005-2021 period. Four out of the eleven identified species were responsible for 86.0% of the infections: Gordonia sputi (53.0%), Gordonia bronchialis (18.3%), Gordonia terrae (8.5%) and Gordonia otitidis (6.1%). Respiratory tract infections (61.6%) and bacteremia (21.9%) were the most common infections. The secA1 gene resolved the inconclusive identification, and two major clonal lineages were observed for G. sputi and G. bronchialis. Species showed a wide antimicrobial susceptibility profile. Cefoxitin resistance varies depending on the species, reaching 94.2% for G. sputi and 36.0% for G. terrae. What is noteworthy is the minocycline resistance in G. sputi (11.5%), the clarithromycin resistance in G. bronchialis secA1 lineage II (30.0%) and the amoxicillin-clavulanate and cefepime resistance in G. terrae (21.4% and 42.8%, respectively). G. sputi and G. bronchialis stand out as the prevalent species causing infections in Spain. Resistance against cefoxitin and other antimicrobials should be considered.

Validity and Reliability of the Short Physical Performance Battery Tool in Institutionalized Spanish Older Adults.

BACKGROUND: In order to be used safely, accurately and reliably, measuring instruments in the health field must first be validated, for which the study of their psychometric properties is necessary. The Short Physical Performance Battery (SPPB) tool is a widely used clinical assessment test that has been approved for usage across several nations, languages and demographics. Finding SPPB's psychometric properties for a sample of institutionalized older individuals is the aim of this research. METHODS: This is a multicenter, retrospective and observational study of the psychometric properties of the Short Physical Performance Battery tool with a convenience sample of 194 institutionalized older adults. Reliability (internal consistency) and validity (construct validity and convergent validity) tests were performed. RESULTS: The results show a very good internal consistency, construct validity and convergent validity. In addition, the factorial structure of the SPPB is provided, which reflects that it is a unidimensional scale. CONCLUSIONS: In conclusion, the Short Physical Performance Battery is a valid and reliable tool for use with institutionalized older adults. Its use is recommended as part of the Comprehensive Geriatric Assessment for the evaluation of the physical or functional sphere. This study was not registered.

National Surveillance of Tetracycline, Erythromycin, and Clindamycin Resistance in Invasive Streptococcus pyogenes: A Retrospective Study of the Situation in Spain, 2007-2020.

BACKGROUND: This work reports on antimicrobial resistance data for invasive Streptococcus pyogenes in Spain, collected by the 'Surveillance Program for Invasive Group A Streptococcus', in 2007-2020. METHODS: emm typing was determined by sequencing. Susceptibility to penicillin, tetracycline, erythromycin, and clindamycin was determined via the E-test. tetM, tetO, msrD, mefA, ermB, ermTR, and ermT were sought by PCR. Macrolide-resistant phenotypes (M, cMLSB, and iMLSB) were detected using the erythromycin-clindamycin double-disk test. Resistant clones were identified via their emm type, multilocus sequence type (ST), resistance genotype, and macrolide resistance phenotype. RESULTS: Penicillin susceptibility was universal. Tetracycline resistance was recorded for 237/1983 isolates (12.0%) (152 carried only tetM, 48 carried only tetO, and 33 carried both). Erythromycin resistance was detected in 172/1983 isolates (8.7%); ermB was present in 83, mefA in 58, msrD in 51, ermTR in 46, and ermT in 36. Clindamycin resistance (methylase-mediated) was present in 78/1983 isolates (3.9%). Eight main resistant clones were identified: two that were tetracycline-resistant only (emm22/ST46/tetM and emm77/ST63/tetO), three that were erythromycin-resistant only (emm4/ST39/mefA-msrD/M, emm12/ST36/mefA-msrD/M, and emm28/ST52/ermB/cMLSB), and three that were tetracycline-erythromycin co-resistant (emm11/ST403/tetM-ermB/cMLSB, emm77/ST63/tetO-ermTR/iMLSB, and emm77/ST63/tetM-tetO-ermTR/iMLSB). CONCLUSIONS: Tetracycline, erythromycin, and clindamycin resistance rates declined between 2007 and 2020. Temporal variations in the proportion of resistant clones determined the change in resistance rates.

Identification and antimicrobial susceptibility of Streptomyces and other unusual Actinobacteria clinical isolates in Spain.

Two hundred and eighty-six isolates from human clinical samples were identified between 1996 and 2019 as belonging to 8 families, 19 genera and 88 species of Actinobacteria. The most identified genera were Streptomyces (182 strains from 45 species), Actinomadura (29 strains, 5 species), Nocardiopsis (21 strains, 6 species) and Dietzia (18 strains, 5 species). The rest of the identified genera (15) contained 27 species with 36 isolates. Of the species studied, only 13/88 had been documented previously as isolates from clinical samples, and in some cases, as true pathogens. In this sense, a literature review of the species found in infections or in clinical samples without clear involvement in pathology has been carried out. Finally, the susceptibility to 8 antimicrobial agents has been studied. Streptomyces showed high resistance (80.8%) against cefotaxime and cotrimoxazole (55.5%), and no isolate resistance to amikacin and linezolid have been found. Lower percentages of resistance have been found in other genera, except in Dietzia (100% against cotrimoxazole and 44.4% against erythromycin). The greatest resistance in these genera was to cotrimoxazole (29.8) and erythromycin (27,9%), and no resistance to linezolid has been found in these genera. In Microbispora, Nonomuraea and Umezawaea, no resistant isolates have been found against any antibiotic studied. Only 3/104 isolates were resistant to amikacin in Amycolatopsis, Crossiella, and Micromonosopora. One isolate of Amycolatopsis was resistant to imipenem.

Invasive Streptococcus pyogenes disease in Spain: a microbiological and epidemiological study covering the period 2007-2019.

The aim of this study is to present the first nationwide microbiological and epidemiological study of invasive group A Streptococcus (iGAS) disease in Spain. One thousand eight hundred ninety-three iGAS isolates were analyzed over 2007-2019. emm typing was performed by sequencing the gene's variable 5' end, exotoxin genes were identified by PCR, and antimicrobial susceptibility explored via the E test and disk diffusion. Five hundred twenty-three isolates were associated with sepsis, 292 with cellulitis, 232 with scarlet fever, 153 with pneumonia, 141 with streptococcal toxic shock syndrome, and 94 with necrotizing fasciitis. The most prevalent emm types were emm1 (449/1893 isolates), emm89 (210/1893), emm3 (208/1893), emm4 (150/1893), emm12 (112/1893) emm6 (107/1893), emm87 (89/1893), emm28 (88/1893), emm75 (78/1893), emm77 (78/1893), emm11 (58/1893), and emm22 (35/1893). emm1, emm3, emm4, and emm6 were the predominant types affecting children (mostly respiratory infections), while emm11, emm77, and emm89 prevailed in the elderly (mostly skin infections). Each emm type was associated with one or more exotoxin gene (spe, sme, and ssa) profiles. speA was detected in 660 isolates, speB in 1829, speC in 1014, speF in 1826, speG in 1651, speJ in 716, speH in 331, smeZ in 720, and ssa in 512. Isolates with speA were associated with the most severe infections. Penicillin susceptibility was universal. Two hundred twenty-four isolates were resistant to tetracycline, 169 to erythromycin, and 81 to clindamycin. Tetracycline, erythromycin, and clindamycin resistance rates declined over the study period. The above information could serve as the basis for continued surveillance efforts designed to control disease cause by this bacterium.

Genomic Background and Phylogeny of cfiA-Positive Bacteroides fragilis Strains Resistant to Meropenem-EDTA.

BACKGROUND: Bacteroides fragilis shows high antimicrobial resistance (AMR) rates and possesses numerous AMR mechanisms. Its carbapenem-resistant strains (metallo-ß-lactamase cfiA-positive) appear as an emergent, evolving clade. METHODS: This work examines the genomes, taxonomy, and phylogenetic relationships with respect to other B. fragilis genomes of two B. fragilis strains (CNM20180471 and CNM20200206) resistant to meropenem+EDTA and other antimicrobial agents. RESULTS: Both strains possessed cfiA genes (cfiA14b and the new cfiA28), along with other AMR mechanisms. The presence of other efflux-pump genes, mexAB/mexJK/mexXY-oprM, acrEF/mdtEF-tolC, and especially cusR, which reduces the entry of carbapenem via the repression of porin OprD, may be related to meropenem-EDTA resistance. None of the detected insertion sequences were located upstream of cfiA. The genomes of these and other B. fragilis strains that clustered together in phylogenetic analyses did not meet the condition of >95% average nucleotide/amino acid identity, or >70% in silico genome-to-genome hybridization similarity, to be deemed members of the same species, although <1% difference in the genomic G+C content was seen with respect to the reference genome B. fragilis NCTC 9343T. CONCLUSIONS: Carbapenem-resistant strains may be considered a distinct clonal entity, and their surveillance is recommended given the ease with which they appear to acquire AMR.

Oxidative and Nitrosative Pattern in Circulating Leukocytes of Very Early/Early Hepatocellular Carcinoma Patients.

BACKGROUND/AIM: In chronic liver disease, various immune cell subsets exert pro or anti-tumour effects by releasing reactive oxygen and nitrogen species (ROS, RNS). Here, we evaluated the oxidative and nitrosative pattern in peripheral blood leukocyte subpopulations of early hepatocellular carcinoma (HCC) patients compared with HCC-free cirrhotic patients. MATERIALS AND METHODS: Venous blood samples from 18 HCC-free cirrhotic patients and 17 early stage HCC patients were collected to determine ROS, RNS and reduced glutathione levels in isolated leukocytes analyzed by flow cytometry. RESULTS: Intracellular levels of ROS and glutathione were higher in lymphocytes, monocytes, and neutrophils from HCC patients as well as mitochondrial superoxide in neutrophils and monocytes whereas intracellular levels of nitric oxide were lower in lymphocytes, monocytes, and neutrophils. CONCLUSION: Early HCC alters intracellular levels of ROS and RNS of some circulating leukocytes subsets. This finding may represent a potential area of interest concerning the development of new treatments and prognostic markers.

Molecular Characterization and Antimicrobial Susceptibilities of Nocardia Species Isolated from the Soil; A Comparison with Species Isolated from Humans.

Nocardia species, one of the most predominant Actinobacteria of the soil microbiota, cause infection in humans following traumatic inoculation or inhalation. The identification, typing, phylogenetic relationship and antimicrobial susceptibilities of 38 soil Nocardia strains from Lara State, Venezuela, were studied by 16S rRNA and gyrB (subunit B of topoisomerase II) genes, multilocus sequence analysis (MLSA), whole-genome sequencing (WGS), and microdilution. The results were compared with those for human strains. Just seven Nocardia species with one or two strains each, except for Nocardia cyriacigeorgica with 29, were identified. MLSA confirmed the species assignments made by 16S rRNA and gyrB analyses (89.5% and 71.0% respectively), and grouped each soil strain with its corresponding reference and clinical strains, except for 19 N. cyriacigeorgica strains found at five locations which grouped into a soil-only cluster. The soil strains of N. cyriacigeorgica showed fewer gyrB haplotypes than the examined human strains (13 vs. 17) but did show a larger number of gyrB SNPs (212 vs. 77). Their susceptibilities to antimicrobials were similar except for beta-lactams, fluoroquinolones, minocycline, and clarithromycin, with the soil strains more susceptible to the first three (p ≤ 0.05). WGS was performed on four strains belonging to the soil-only cluster and on two outside it, and the results compared with public N. cyriacigeorgica genomes. The average nucleotide/amino acid identity, in silico genome-to-genome hybridization similarity, and the difference in the genomic GC content, suggest that some strains of the soil-only cluster may belong to a novel subspecies or even a new species (proposed name Nocardia venezuelensis).

Saezia sanguinis gen. nov., sp. nov., a Betaproteobacteria member of order Burkholderiales, isolated from human blood.

The taxonomic position of an unknown bacterial strain designated CNM695-12, isolated from the blood of an immunocompromised subject, was investigated via phenotypic, chemotaxonomic, genotypic and genomic analyses. Bacterial cells were determined to be Gram-stain-negative bacilli, aerobic, non-motile and non-spore-forming. The strain showed catalase activity but no oxidase activity. Optimal growth occurred at 37 °C, pH 7 and with 0-1 % NaCl. C16 : 0, summed feature 8 (comprising C18 : 1ω7c /C18:1 ω6c), and C18 : 1ω9c were the most abundant fatty acids, and ubiquinone 8 was the major respiratory quinone. The polar lipids present included phosphatidylglycerol, phosphatidylethanolamine and other aminophospholipids. The 16S rRNA gene sequence showed approximately 93.5 % similarity to those of different species with validly published names within the order Burkholderiales (e.g. Leptothrix mobilis Feox-1T, Aquabacterium commune B8T , Aquabacterium citratiphilum B4T and Schlegelella thermodepolymerans K14T). Phylogenetic analyses based on 16S rRNA gene sequences and concatenated alignments including the sequences for 107 essential proteins, revealed the strain to form a novel lineage close to members of the family Comamonadaceae. The highest average nucleotide identity and average amino acid identity values were obtained with Schlegelella thermodepolymerans K14T (69.6 and 55.7 % respectively). The genome, with a size of 3.35 Mb, had a DNA G+C content of 52.4 mol% and encoded 3056 predicted genes, 3 rRNA, 1 transfer-messengerRNA and 51 tRNA. Strain CNM695-12 thus represents a novel species belonging to a novel genus within the order Burkholderiales, for which the name Saezia sanguinis gen. nov., sp. nov. is proposed. The type strain is CNM695-12T (=DSM 104959T=CECT 9208T).

Dynamics of a Sporadic Nosocomial Acinetobacter calcoaceticus - Acinetobacter baumannii Complex Population.

Our objective was to improve current knowledge of sporadic (Spo) nosocomial Acinetobacter calcoaceticus-Acinetobacter baumannii (Acb) complex populations, and thus better understand the epidemiology of Spo and endemoepidemic (EE) strains. Between 1999 and 2010, 133 isolates of Spo Acb complex were obtained from a single hospital. Species were identified by gyrB-PCR, and via gyrB- and rpoB-sequencing. Clonal analysis was undertaken using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. Susceptibility to antimicrobial agents was determined by microdilution and E-tests. Carbapenemase genes were detected by PCR. One hundred and one PFGE types were detected. A. baumannii was the most common (67/101 PFGE types), followed by Acinetobacter pittii (22/101), Acinetobacter lactucae (6/101), and Acinetobacter calcoaceticus (2/101). gyrB, rpoB1, and rpoB2 sequencing returned 49, 13, and 16 novel sequences, respectively. Sixty-three sequence types (STs) (38 new STs and 66 new alleles) were detected; the most common were ST2 (29/133 isolates) and ST132 (14/133). Twenty-six OXA-51 allelic variants were detected, nine of which were novel. The PFGE types were generally susceptible (88/101) to all the tested antimicrobials; 3/101 were carbapenem-resistant due to the presence of the genetic structure ISAba2-bla OXA-58-like-ISAba3, and 2/101 were multidrug-resistant. It can be concluded that the examined Spo Acb complex population was mainly composed of A. baumannii. Many different clones were detected (with ST2 clearly dominant), all largely susceptible to antimicrobials; multidrug resistance was rare. In contrast, a previously examined EE Acb population was composed of just four expanding, multidrug-resistant A. baumannii clones -ST2, ST3, ST15, and ST80-.

First Insight into the Genome Sequences of Two Linezolid-Resistant Nocardia farcinica Strains Isolated from Patients with Cystic Fibrosis.

The draft genome sequences of two Nocardia farcinica strains isolated from two patients with cystic fibrosis (CF), resistant to trimethoprim/sulfamethoxazole and linezolid, are reported here. The estimated genome sizes were 5.8 Mb with a 70.63% G+C content. Transposases from Tn916 were detected, but not 23S rRNA mutation (G2576T) related to linezolid resistance.

Paenibacillus spp. isolated from human and environmental samples in Spain: detection of 11 new species.

One hundred thirty-six isolates, 88 human and 48 environmental, that met the requirements to belong to the genus Paenibacillus were identified using a polyphasic taxonomic approach known as 16S rRNA plus phenotypic traits. Thirty-seven Paenibacillus species were identified; some had not been previously reported from clinical samples. The main species were P. pabuli (13 isolates), P. provencensis (11), P. phoenicis (9) and P. lautus (8). P. pabuli (11/13) and P. provencensis (8/11) were mainly environmental isolates, while P. phoenicis (9/9) and P. lautus (6/8) were mainly human isolates. Despite the difficulties in assigning to human Paenibacillus isolates a role as a pathogen or contaminant, here 25% of the isolates were involved in true infections, especially in those cases that affected abscesses, wound exudates, ocular infections and diverse fluids. In addition, 15 isolates were identified as 11 'Candidatus' to a new species, all of them from human specimens except one that was obtained from laboratory air. The antimicrobial susceptibility testing showed 95.6% of isolates were resistant to ampicillin, 44% were resistant to cotrimoxazole, 20 to 30% were resistant to cefotaxime and vancomycin and 13% were resistant to rifampicin and erythromycin.

Epidemiology and susceptibility to antimicrobial agents of the main Nocardia species in Spain.

Objectives: The aims of this study were to explore the clinical distribution, by species, of the genus Nocardia and to assess the antimicrobial susceptibilities of the 10 most prevalent species identified in Spain. Methods: Over a 10 year period (2005-14), 1119 Nocardia strains were molecularly identified and subjected to the Etest. The distribution and resistance trends over the sub-periods 2005-09 and 2010-14 were also examined. Results: Of the strains examined, 82.9% belonged to the following species: Nocardia cyriacigeorgica (25.3%), Nocardia nova (15.0%), Nocardia abscessus (12.7%), Nocardia farcinica (11.4%), Nocardia carnea (4.3%), Nocardia brasiliensis (3.5%), Nocardia otitidiscaviarum (3.1%), Nocardia flavorosea (2.6%), Nocardia rhamnosiphila (2.6%) and Nocardia transvalensis (2.4%). Their prevalence values were similar during 2005-09 and 2010-14, except for those of N. abscessus , N. farcinica and N. transvalensis , which fell significantly in the second sub-period ( P ≤ 0.05). The major location of isolation was the respiratory tract (∼86%). Half (13/27) of all strains from the CNS were N. farcinica . Significant differences in MIC results were recorded for some species between the two sub-periods. According to the CLSI's breakpoints, low resistance rates (≤15%) were recorded for seven species with respect to cefotaxime, imipenem and tobramycin; five species showed similar rates with respect to trimethoprim/sulfamethoxazole. Linezolid and amikacin were the most frequently active agents. Conclusion: The accurate identification of the infecting species and the determination of its susceptibility to antimicrobial agents, given the large number of strains with atypical patterns, are crucial if patients with nocardiosis are to be successfully treated.

Lactational mastitis caused by Streptococcus lactarius.

Human infections caused by Streptococcus lactarius have not been previously reported. In the present report, we describe a lactational mastitis caused by this organism. The infection occurred in a 28-year-old breast-feeding female, with a 10-days history of moderate pain on the right breast. The patient was cured after antibiotic treatment with levofloxacin for 21 days. Our case shows that S. lactarius should be considered as a cause of lactational mastitis. The introduction of molecular microbiology techniques can be extremely useful for knowing the implication of streptococci in lactational mastitis.

Decreased bioavailability of nitric oxide in aorta from ovariectomized senescent mice. Role of cyclooxygenase.

This study investigates the effects of aging and/or ovariectomy on vascular reactivity to thromboxane A2 (TXA2) receptor stimulation with U46619, and the modulation by nitric oxide (NO) and cyclooxygenase (COX) in aorta from female senescence-accelerated mice (SAMP8) and from senescence resistant mice (SAMR1). Five-month-old female SAMR1 and SAMP8 were divided into three groups: sham-operated, ovariectomized and ovariectomized plus estradiol. Twenty-eight days after surgery, thoracic aortic rings were mounted for isometric recording of tension and concentration-response curves for U46619 (10(-10)-3 × 10(-7) M) were performed in the absence and in the presence of the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME, 10(-4) M) and/or COX inhibitor indomethacin (10(-5)M). Vascular superoxide production was detected by dihydroethidium staining on sections of thoracic aorta. NO bioavailability in response to U46619 was suppressed by estrogen withdrawn in young and senescent mice and was restored by the administration of estradiol. In the presence of indomethacin, contractions to U46619 decreased in all groups indicating an aging- and estrogen-dependent modulation of contractile prostanoids. The simultaneous incubation of L-NAME and indomethacin did not change the maximal responses and sensitivities to TXA2 in any group in comparison with untreated aortic segments. The superoxide generation induced by TXA2 was greater in aorta from SAMP8 than in SAMR1. Moreover, in ovariectomized groups superoxide production was further increased and treatment with 17ß-estradiol reverted the effects of the ovariectomy. Inhibition of COX with indomethacin prevented the U46619-induced increase in superoxide formation. Our results indicate that NO bioavailability in response to TP receptor activation is both estrogen- and aging-dependent. TXA2 induced contractions are partially mediated by COX activation. Both aging and ovariectomy enhanced COX-dependent component of the TXA2-induced contraction. It is noteworthy that in the absence of estrogen, COX inhibition induces an increase of NO bioavailability. Therefore, in senescent female mice with an experimental menopause, TP-receptor stimulation is responsible for COX activation and enhanced superoxide generation, which may result in reduced NO bioavailability. These effects were reversed by estrogen administration.

Effects of asymmetric dimethylarginine on renal arteries in portal hypertension and cirrhosis.

AIM: To evaluate the effects of asymmetric dimethylarginine (ADMA) in renal arteries from portal hypertensive and cirrhotic rats. METHODS: Rat renal arteries from Sham (n = 15), pre-hepatic portal hypertension (PPVL; n = 15) and bile duct ligation and excision-induced cirrhosis (BDL; n = 15) were precontracted with norepinephrine, and additional contractions were induced with ADMA (10-6-10-3 mol/L), an endogenous inhibitor of nitric oxide (NO) synthase. Concentration-response curves to acetylcholine (1 × 10-9-3 × 10-6 mol/L) were determined in precontracted renal artery segments with norepinephrine in the absence and in the presence of ADMA. Kidneys were collected to determine the protein expression and activity of dimethylarginine dimethylaminohydrolase (DDAH), an enzyme that catabolizes ADMA. RESULTS: In renal arteries precontracted with norepinephrine, ADMA caused endothelium-dependent contractions. The pD2 values to ADMA were similar in the Sham and PPVL groups (4.20 ± 0.08 and 4.11 ± 0.09, P > 0.05, respectively), but were lower than those of the BDL group (4.79 ± 0.16, P < 0.05). Acetylcholine-induced endothelium-dependent relaxation that did not differ, in terms of pD2 and maximal relaxation, among the 3 groups studied. Treatment with ADMA (3 × 10-4 mol/L) inhibited acetylcholine-induced relaxation in the 3 groups, but the inhibition was higher (P < 0.05) in the BDL group compared with that for the Sham and PPVL groups. The mRNA and protein expression of DDAH-1 were similar in kidneys from the three groups. Conversely, DDAH-2 expression was increased (P < 0.05) in PPVL and further enhanced (P < 0.05) in the BDL group. However, renal DDAH activity was significantly decreased in the BDL group. CONCLUSION: Cirrhosis increased the inhibitory effect of ADMA on basal- and induced-release of NO in renal arteries, and decreased DDAH activity in the kidney.

Asymmetric dimethylarginine as a mediator of vascular dysfunction in cirrhosis.

Cirrhosis is associated with marked abnormalities in the circulatory function that involve a reduction in systemic vascular resistance. An important cause of this vasodilatation is the increased production or activity of nitric oxide (NO) in the splanchnic circulation. During portal hypertension and cirrhosis an increased endothelial NO synthase (eNOS) activity is demonstrated in splanchnic vessels. In contrast, the activity of eNOS in the cirrhotic liver is decreased, which suggests a different regulation of eNOS in the liver and in the splanchnic vessels. Asymmetric dimethylarginine (ADMA) is an endogenous NO inhibitor and higher plasma levels of ADMA are related to increased cardiovascular risk in both the general population and among patients with cirrhosis. It has been demonstrated that the liver is a key player in the metabolism of ADMA. This observation was further supported by investigations in human patients, showing a close correlation between ADMA plasma levels and the degree of hepatic dysfunction. ADMA is degraded to citrulline and dimethylamine by dimethylarginine dimethylaminohydrolases (DDAHs). DDAHs are expressed as type 1 and 2 isoforms and are widely distributed in various organs and tissues, including the liver. In this review, we discuss experimental and clinical data that document the effects of dimethylarginines on vascular function in cirrhosis. Our increasing understanding of the routes of synthesis and metabolism of methylarginines is beginning to provide insights into novel mechanisms of liver disease and allowing us to identify potential therapeutic opportunities.

Resistance gene pool to co-trimoxazole in non-susceptible Nocardia strains.

The soil-borne pathogen Nocardia sp. causes severe cutaneous, pulmonary, and central nervous system infections. Against them, co-trimoxazole (SXT) constitutes the mainstay of antimicrobial therapy. However, some Nocardia strains show resistance to SXT, but the underlying genetic basis is unknown. We investigated the presence of genetic resistance determinants and class 1-3 integrons in 76 SXT-resistant Nocardia strains by PCR and sequencing. By E test, these clinical strains showed SXT minimum inhibitory concentrations of ≥32:608 mg/L (ratio of 1:19 for trimethoprim: sulfamethoxazole). They belonged to 12 species, being the main representatives Nocardia farcinica (32%), followed by N. flavorosea (6.5%), N. nova (11.8%), N. carnea (10.5%), N. transvalensis (10.5%), and Nocardia sp. (6.5%). The prevalence of resistance genes in the SXT-resistant strains was as follows: sul1 and sul2 93.4 and 78.9%, respectively, dfrA(S1) 14.7%, blaTEM-1 and blaZ 2.6 and 2.6%, respectively, VIM-2 1.3%, aph(3')-IIIa 40.8%, ermA, ermB, mefA, and msrD 2.6, 77.6, 14.4, and 5.2%, respectively, and tet(O), tet(M), and tet(L) 48.6, 25.0, and 3.9%, respectively. Detected amino acid changes in GyrA were not related to fluoroquinolone resistance, but probably linked to species polymorphism. Class 1 and 3 integrons were found in 93.42 and 56.57% strains, respectively. Class 2 integrons and sul3 genes were not detected. Other mechanisms, different than dfrA(S1), dfrD, dfrF, dfrG, and dfrK, could explain the strong trimethoprim resistance shown by the other 64 strains. For first time, resistance determinants commonly found in clinically important bacteria were detected in Nocardia sp. sul1, sul2, erm(B), and tet(O) were the most prevalent in the SXT-resistant strains. The similarity in their resistome could be due to a common genetic platform, in which these determinants are co-transferred.