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1.
Eur J Clin Microbiol Infect Dis ; 40(11): 2295-2303, 2021 Nov.
Article in English | MEDLINE | ID: mdl-34046804

ABSTRACT

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.


Subject(s)
Streptococcal Infections/microbiology , Streptococcus pyogenes/isolation & purification , Adolescent , Adult , Aged , Aged, 80 and over , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Carrier Proteins/genetics , Carrier Proteins/metabolism , Child , Child, Preschool , Erythromycin/pharmacology , Exotoxins/genetics , Exotoxins/metabolism , Female , Humans , Infant , Male , Membrane Proteins/genetics , Membrane Proteins/metabolism , Microbial Sensitivity Tests , Middle Aged , Penicillins/pharmacology , Spain/epidemiology , Streptococcal Infections/epidemiology , Streptococcus pyogenes/classification , Streptococcus pyogenes/drug effects , Streptococcus pyogenes/genetics , Young Adult
2.
Front Microbiol ; 10: 593, 2019.
Article in English | MEDLINE | ID: mdl-30967856

ABSTRACT

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-.

3.
J Antimicrob Chemother ; 72(3): 754-761, 2017 03 01.
Article in English | MEDLINE | ID: mdl-27999029

ABSTRACT

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.


Subject(s)
Anti-Bacterial Agents/pharmacology , Nocardia Infections/microbiology , Nocardia/drug effects , Amikacin/therapeutic use , Anti-Bacterial Agents/therapeutic use , Humans , Imipenem/therapeutic use , Linezolid/pharmacology , Microbial Sensitivity Tests , Nocardia/classification , Nocardia/genetics , Nocardia/isolation & purification , Nocardia Infections/drug therapy , Nocardia Infections/epidemiology , Prevalence , RNA, Ribosomal, 16S , Spain/epidemiology , Trimethoprim, Sulfamethoxazole Drug Combination/pharmacology , Trimethoprim, Sulfamethoxazole Drug Combination/therapeutic use
4.
Front Microbiol ; 7: 542, 2016.
Article in English | MEDLINE | ID: mdl-27148228

ABSTRACT

Nocardia species are difficult to identify, a consequence of the ever increasing number of species known and their homogeneous genetic characteristics. 16S rRNA analysis has been the gold standard for identifying these organisms, but proteomic techniques such as matrix-assisted laser desorption ionization-time of flight (MALDI-TOF MS) and housekeeping gene analysis, have also been explored. One hundred high (n = 25), intermediate (n = 20), and low (n = 55) prevalence (for Spain) Nocardia strains belonging to 30 species were identified via 16S rRNA and MALDI-TOF MS analysis. The manufacturer-provided database MALDI Biotyper library v4.0 (5.627 entries, Bruker Daltonik) was employed. In the high prevalence group (Nocardia farcinica, N. abscessus, N. cyriacigeorgica and N. nova), the 16S rRNA and MALDI-TOF MS methods provided the same identification for 76% of the strains examined. For the intermediate prevalence group (N. brasiliensis, N. carnea, N. otitidiscaviarum and N. transvalensis complex), this figure fell to 45%. In the low-prevalence group (22 species), these two methods were concordant only in six strains at the species level. Tetra-gene multi-locus sequencing analysis (MLSA) involving the concatemer gyrB-16S rRNA-hsp65-secA1 was used to arbitrate between discrepant identifications (n = 67). Overall, the MLSA confirmed the results provided at species level by 16S rRNA analysis in 34.3% of discrepancies, and those provided by MALDI-TOF MS in 13.4%. MALDI-TOF MS could be a strong candidate for the identification of Nocardia species, but only if its reference spectrum database improves, especially with respect to unusual, recently described species and species included in the described Nocardia complexes.

5.
Front Microbiol ; 6: 376, 2015.
Article in English | MEDLINE | ID: mdl-25972856

ABSTRACT

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.

6.
BMC Microbiol ; 15: 47, 2015 Feb 25.
Article in English | MEDLINE | ID: mdl-25887224

ABSTRACT

BACKGROUND: Nosocomial outbreaks of multidrug-resistant Acinetobacter baumannii are of worldwide concern. Using pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and multiple locus variable number tandem repeat sequence (VNTR) analysis (MLVA), the present work examines the genetic diversity of the endemic and epidemic A. baumannii clones isolated in a single hospital over a twelve-year period. RESULTS: PFGE analysis of 405 A. baumannii-calcoaceticus complex isolates detected 15 A. baumannii endemic/epidemic PFGE types (EE1 to EE15) that grouped into five clusters: EE1-EE8, EE9, EE10, EE11 and EE12-EE15. The MLST sequence type (ST) distributions were: international clone II (ST-2) 60%, international clone III (ST-3) 26.7%, ST-15 6.7%, and ST-80 6.7%. MLVA-8Orsay returned 17 allelic profiles. The large (L) VNTR marker profiles were fully concordant with the detected STs, and concordant with 14 up to 15 PFGE types. Imipenem resistance was detected in five PFGE types; the prevalence of the bla OXA-58-like and bla OXA-40-like genes was 60% and 40% respectively. CONCLUSIONS: PFGE proved to be a vital tool for analysis of the temporal and spatial distribution of the clones. MLST and the VNTR L-markers grouped the isolates into clonal clusters. The wide diversity of MLVA small (S)-markers, however, did not permit clustering. The present results demonstrate the persistence of several endemic PFGE types in the hospital, the involvement of some of them in outbreaks, and the inter hospital transmission of extensively drug-resistant ST-15 and ST-80.


Subject(s)
Acinetobacter Infections/epidemiology , Acinetobacter Infections/microbiology , Acinetobacter baumannii/classification , Acinetobacter baumannii/isolation & purification , Cross Infection/epidemiology , Cross Infection/microbiology , Genotype , Acinetobacter baumannii/genetics , DNA, Bacterial/genetics , Humans , Minisatellite Repeats , Molecular Epidemiology , Multilocus Sequence Typing , Polymorphism, Restriction Fragment Length , Tertiary Care Centers
7.
J Clin Microbiol ; 53(3): 997-1001, 2015 Mar.
Article in English | MEDLINE | ID: mdl-25540402

ABSTRACT

gyrB is used to improve the identification of the Nocardia species N. brasiliensis, N. higoensis, N. ignorata, N. otitidiscaviarum, N. paucivorans, N. pneumoniae, N. puris, N. takedensis, N. veterana, and N. vinacea, but it does not improve the identification of another 12 Nocardia studied species. gyrB provides typing and phylogenetic markers for N. carnea, N. transvalensis, N. brasiliensis, and N. otitidiscaviarum.


Subject(s)
DNA Gyrase/genetics , Nocardia/classification , Nocardia/isolation & purification , Phylogeny , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Genetic Markers , Humans , Molecular Sequence Data , Molecular Typing , Nocardia/genetics , Sequence Analysis, DNA
9.
J Clin Microbiol ; 51(11): 3602-8, 2013 Nov.
Article in English | MEDLINE | ID: mdl-23966490

ABSTRACT

This study compares the identification, typing, and phylogenetic relationships of the most prevalent clinical Nocardia species in Spain, as determined via sequence analysis of their housekeeping genes gyrB and rpoB, with the results returned by the gold standard 16S rRNA method. gyrB and rpoB analyses identified Nocardia abscessus, N. cyriacigeorgica, N. farcinica, and the N. nova complex, species that together account for more than half of the human nocardiosis cases recorded in Spain. The individual discriminatory power of gyrB and rpoB with respect to intraspecies typing, calculated using the Hunter-Gaston discriminatory index (HGDI), was generally high (HGDI, 0.85 to 1), except for rpoB with respect to N. farcinica (HGDI, 0.71). Phylogenetically, different degrees of intra- and interspecies microheterogeneity were observed for gyrB and rpoB in a group of 119 clinical strains. A single 16S haplotype was obtained for each species, except for the N. nova complex (8 types), while gyrB and rpoB were more polymorphic: N. abscessus had 14 and 18 haplotypes, N. cyriacigeorgica had 17 and 12, N. farcinica had 11 and 5, and the N. nova complex had 26 and 29 haplotypes, respectively. A diversity gradient was therefore seen, with N. farcinica at the bottom followed by N. abscessus and N. cyriacigeorgica in the middle and N. nova complex at the top. The complexity of the N. nova complex is highlighted by its six variations in the GyrB (126)AAAPEH motif. gyrB sequencing (with or without rpoB sequencing) offers a simple means for identifying the most prevalent Nocardia species in Spanish medical laboratories and for determining the intraspecific diversity among their strains.


Subject(s)
DNA Gyrase/genetics , DNA-Directed RNA Polymerases/genetics , Genetic Variation , Nocardia Infections/microbiology , Nocardia/classification , Nocardia/isolation & purification , RNA, Ribosomal, 16S/genetics , Cluster Analysis , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , DNA, Ribosomal/chemistry , DNA, Ribosomal/genetics , Humans , Molecular Epidemiology , Molecular Sequence Data , Nocardia/genetics , Nocardia Infections/epidemiology , Phylogeny , Prevalence , Sequence Analysis, DNA , Spain/epidemiology
10.
J Antimicrob Chemother ; 68(3): 550-3, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23143900

ABSTRACT

OBJECTIVES: To study the distribution, diversity and activity of Acinetobacter-derived cephalosporinase (ADC)-, carbapenem-hydrolysing oxacillinase (CHO)- and metallo-ß-lactamase (MBL)-encoding genes, and of the most common insertion sequences (ISs), in the genome of nosocomial, epidemic, multidrug-resistant Acinetobacter baumannii (MDRAB) clones from Spain. METHODS: The studied population included 59 MDRAB strains previously genotyped by PFGE and multilocus sequence typing. The search for the ADC (bla(ADC)), CHO (bla(OXA-51-like), bla(OXA-23-like), bla(OXA-40-like) and bla(OXA-58-like)) and MBL (bla(IMP), bla(VIM), bla(SIM-1), bla(GIM-1), bla(SPM-1) and bla(NDM-1)) genes, and for the ISs (ISAba1, ISAba2, ISAba3, ISAba4 and IS18) was done by PCR assays. The phenotypic presence of MBL enzymes was examined using imipenem/imipenem + EDTA strips. RESULTS: The most prevalent IS, ISAba1 (93.2%), was detected upstream of bla(ADC) and bla(OXA-51-like). These genes showed ample diversity (10 and 8 alleles, respectively). Four ADC sequences (ADC-1-like(P240S), ADC-2-like(N260H/T264N), ADC-11-like(Q163K) and ADC-11-like(G342R)) are described here for the first time. bla(OXA-58-like) was carried by 20.3% of strains, in association with ISAba2, ISAba3 or IS18. bla(OXA-40-like) was the most prevalent acquired CHO gene (57.6%), and was associated with none of the studied ISs. Neither bla(OXA-23-like) nor ISAba4 was detected in any strain. Some 67.8% of strains with MBL activity showed no corresponding gene in PCR; these results were more common in strains with a highly active CHO, such as OXA-40. CONCLUSIONS: All the studied genes and their related ISs showed a clonal distribution. Imipenem resistance was probably provided by OXA-40 for the most part, while MBL- and OXA-23-encoding genes were absent in the studied population.


Subject(s)
Acinetobacter Infections/epidemiology , Acinetobacter baumannii/enzymology , DNA Transposable Elements , Genetic Variation , beta-Lactamases/genetics , Acinetobacter Infections/microbiology , Acinetobacter baumannii/genetics , Acinetobacter baumannii/isolation & purification , Cluster Analysis , Cross Infection/epidemiology , Cross Infection/microbiology , DNA, Bacterial/genetics , Drug Resistance, Multiple, Bacterial , Genotype , Humans , Microbial Sensitivity Tests , Molecular Epidemiology , Molecular Typing , Polymerase Chain Reaction , Spain/epidemiology
12.
J Antimicrob Chemother ; 67(4): 832-6, 2012 Apr.
Article in English | MEDLINE | ID: mdl-22228676

ABSTRACT

OBJECTIVES: To report a clonal outbreak of ST17 vancomycin-resistant Enterococcus faecium (VREfm) carrying Tn1546 (vanA) in a haemo-oncology ward of a tertiary teaching hospital in the south of Spain (January-September 2009). PATIENTS AND METHODS: Twenty-two VREfm strains from 13 patients were characterized by PFGE, multiple-locus variable-number tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). Genes encoding antibiotic resistance and putative virulence traits and the Tn1546 backbone were investigated by PCR. Plasmid characterization included determination of size (S1-PFGE) and replication modules (PCR, hybridization and sequencing). Patient clinical records were analysed retrospectively. RESULTS: A single ST17 E. faecium clone (MT-7 MLVA type) carrying esp and hyl plus a 30 kb Inc18-like::Tn1546 (IS1216) plasmid was identified. Ampicillin resistance was linked to PBP5 showing mutations at positions 24, 27, 34, 66, 68, 85, 100, 144, 172, 177, 204, 216, 324, 462, 466', 470, 485, 496, 499, 525, 546, 558, 582, 586, 629, 632, 642 and 667. Other resistance genes identified were erm(B), ant(6')-Ia and aph(3')-IIIa. Fluoroquinolone resistance was attributable to ParC (Arg-61 → Gly and Ser-80 → Arg) and GyrA (Ser-83 → Arg) mutations. CONCLUSIONS: A nosocomial outbreak caused by an ST17 (CC17) E. faecium clone harbouring Esp and Hyl and a 30 kb Inc18-like::Tn1546 plasmid among haemo-oncology patients is reported. The failure of early infection control practices indicates an undetected reservoir and the ability of this strain to persist over long periods. The potential spread of epidemic clones and broad host plasmids carrying vancomycin resistance in Spain is of concern since it might contribute towards a higher rate of VREfm infection.


Subject(s)
Cross Infection/epidemiology , DNA Transposable Elements , Disease Outbreaks , Enterococcus faecium/isolation & purification , Gram-Positive Bacterial Infections/epidemiology , Hematologic Neoplasms/complications , Plasmids , Adult , Cluster Analysis , Cross Infection/microbiology , Electrophoresis, Gel, Pulsed-Field , Enterococcus faecium/classification , Enterococcus faecium/genetics , Female , Gram-Positive Bacterial Infections/microbiology , Hospitals , Humans , Male , Middle Aged , Minisatellite Repeats , Molecular Sequence Data , Molecular Typing , Polymerase Chain Reaction , Sequence Analysis, DNA , Spain/epidemiology , Vancomycin Resistance , Virulence Factors/genetics
13.
J Clin Microbiol ; 49(3): 875-82, 2011 Mar.
Article in English | MEDLINE | ID: mdl-21177889

ABSTRACT

Acinetobacter baumannii is one of the major pathogens involved in nosocomial outbreaks. The clonal diversity of 729 epidemic strains isolated from 19 Spanish hospitals (mainly from intensive care units) was analyzed over an 11-year period. Pulsed-field gel electrophoresis (PFGE) identified 58 PFGE types that were subjected to susceptibility testing, rpoB gene sequencing, and multilocus sequence typing (MLST). All PFGE types were multidrug resistant; colistin was the only agent to which all pathogens were susceptible. The 58 PFGE types were grouped into 16 clones based on their genetic similarity (cutoff of 80%). These clones were distributed into one major cluster (cluster D), three medium clusters (clusters A, B, and C), and three minor clusters (clusters E, F, and G). The rpoB gene sequencing and MLST results reflected a clonal distribution, in agreement with the PFGE results. The MLST sequence types (STs) (and their percent distributions) were as follows: ST-2 (47.5%), ST-3 (5.1%), ST-15 (1.7%), ST-32 (1.7%), ST-79 (13.6%), ST-80 (20.3%), and ST-81 (10.2%). ST-79, ST-80, and ST-81 and the alleles cpn60-26 and recA29 are described for the first time. International clones I, II, and III were represented by ST-81, ST-2, and ST-3, respectively. ST-79 and ST-80 could be novel emerging clones. This work confirms PFGE and MLST to be complementary tools in clonality studies. Here PFGE was able to demonstrate the monoclonal pattern of most outbreaks, the inter- and intrahospital transmission of bacteria, and their endemic persistence in some wards. MLST allowed the temporal evolution and spatial distribution of Spanish clones to be monitored and permitted international comparisons to be made.


Subject(s)
Acinetobacter Infections/epidemiology , Acinetobacter baumannii/classification , Acinetobacter baumannii/genetics , Bacterial Typing Techniques , Cross Infection/epidemiology , Molecular Typing , Polymorphism, Genetic , Acinetobacter Infections/microbiology , Acinetobacter baumannii/isolation & purification , Cluster Analysis , Cross Infection/microbiology , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , DNA-Directed RNA Polymerases/genetics , Electrophoresis, Gel, Pulsed-Field , Genotype , Humans , Molecular Epidemiology , Molecular Sequence Data , Multilocus Sequence Typing , Sequence Analysis, DNA , Spain/epidemiology
14.
J Clin Microbiol ; 48(8): 2734-40, 2010 Aug.
Article in English | MEDLINE | ID: mdl-20554816

ABSTRACT

The severe morbidity of human brucellosis is one of the main reasons for using molecular typing in the epidemiological surveillance of this worldwide zoonosis. Multiple-locus variable-number repeat analysis (MLVA-16), hypervariable octameric oligonucleotide fingerprinting (HOOF-print), and the differences in the single nucleotide polymorphisms (SNPs) (codons 1249 and 1309) of the DNA-dependent RNA polymerase beta subunit (rpoB) were used to type a human Brucella melitensis population (108 strains) collected from throughout Spain over 13 years. Eighty-six MLVA types (discriminatory index, 0.99) were detected, with a wide-ranging genetic similarity coefficient (37.2 to 93.7%). The population clustered into the following groups: American, with genotypes 47 (1 strain), 48 (13 strains), 53 (12 strains), 55 (2 strains), 80 (1 strain), and a new genotype (2 strains), Western Mediterranean, with genotype 51 (9 strains), and Eastern Mediterranean, with genotypes 42 (60 strains), 43 (4 strains), and 63 (4 strains). Two profession-related and two foodborne acquisitions were confirmed. Distributed throughout Spain, Eastern Mediterranean genotype 42 was the most common (55%). The low MLVA-16 allelic polymorphism (genetic similarity range, 75 to 94%) of the genotype 42 strains suggests that they recently evolved from a common ancestor. rpoB typing grouped the strains as rpoB type 1 (1249-ATG/1309-CTG; 28.7%), rpoB type 2 (1249-ATG/1309-CTA; 62.9%), and rpoB type 3 (1249-ATA/1309-CTG; 8.3%). According to the MLVA-16 results, the population clustered by rpoB type. Given the correlation between B. melitensis MLVA groups and rpoB types (American and rpoB type 1, Eastern Mediterranean and rpoB type 2, and Western Mediterranean and rpoB type 3), the rpoB type could be used as an initial marker for the epidemiological surveillance of brucellosis.


Subject(s)
Bacterial Typing Techniques , Brucella melitensis/classification , Brucella melitensis/isolation & purification , Brucellosis/epidemiology , Brucellosis/microbiology , DNA Fingerprinting , DNA, Bacterial/genetics , Bacterial Proteins/genetics , Brucella melitensis/genetics , Cluster Analysis , DNA-Directed RNA Polymerases/genetics , Genotype , Humans , Minisatellite Repeats , Molecular Epidemiology , Oligonucleotides/genetics , Polymorphism, Genetic , Spain/epidemiology
16.
J Antimicrob Chemother ; 65(1): 51-3, 2010 Jan.
Article in English | MEDLINE | ID: mdl-19861338

ABSTRACT

OBJECTIVES: The aim of this study was to determine, using molecular methods, whether rifampicin and fluoroquinolone resistance was present in a clinical Brucella melitensis population. METHODS: Sixty-two B. melitensis strains, isolated from humans-most experiencing their first brucellosis episode-over an 11 year period in Spain, were genotyped by multiple locus variable analysis (MLVA-16) for future studies. In the present work, molecular screening was undertaken to detect the presence of rpoB and gyrA/gyrB/parC/parE mutations (previously described in in vitro Brucella spp. mutants) related to resistance to rifampicin and fluoroquinolones, respectively. RESULTS: Sixty-two MLVA-16 genotypes were identified among the B. melitensis population, with genetic similarity values ranging from 32% to 94%. rpoB mutations related to rifampicin resistance (positions 154, 526, 536, 539, 541, 574) were not detected. Neither were changes in GyrA described in in vitro mutants (67, 71, 87, 91 and an insertion at 340) detected in these strains. All showed identical GyrA, GyrB, ParC and ParE sequences with respect to B. melitensis 16M, except for one strain (ciprofloxacin and moxifloxacin MICs 0.25-0.50 mg/L) that harboured the Val264Ala replacement outside the GyrA quinolone resistance-determining region (QRDR); no differences were seen, however, in the NorMI/II efflux pump genes. CONCLUSIONS: The absence of rpoB mutations clearly related to rifampicin resistance in clinical B. melitensis strains reinforces the first-choice status of this antibiotic in the treatment of first brucellosis episodes, and demonstrates the usefulness of molecular screening for resistant genotypes. The absence of topoisomerase II-IV mutations, however, cannot rule out fluoroquinolone resistance due to the interplay of different mechanisms.


Subject(s)
Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Brucella melitensis/drug effects , Brucella melitensis/genetics , DNA, Bacterial/genetics , Drug Resistance, Bacterial , Fluoroquinolones/pharmacology , Rifampin/pharmacology , Brucella melitensis/isolation & purification , Brucellosis/microbiology , Humans , Mass Screening , Molecular Sequence Data , Mutation, Missense , Point Mutation , Sequence Analysis, DNA , Spain
18.
J Antimicrob Chemother ; 63(1): 17-20, 2009 Jan.
Article in English | MEDLINE | ID: mdl-19001448

ABSTRACT

OBJECTIVES: A large clonal outbreak of multidrug-resistant CC17 ST17 Enterococcus faecium containing Tn5382 in a hospital in the north of Spain is described. METHODS: We characterized vancomycin-resistant E. faecium isolates from 10 infected and 40 colonized inpatients from a single hospital by PFGE, multiple-locus variable-number tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). Genes encoding antibiotic resistance (ampicillin, aminoglycosides, macrolides, quinupristin/dalfopristin, quinolones, tetracycline) and putative virulence traits were analysed. RESULTS: All isolates showed highly similar PFGE profiles and were assigned to the type MT1 by MLVA and to ST17 (CC17) by MLST. The Tn5382 type identified in all isolates was linked to pbp5 and contained a 5 bp deletion and 10 point mutations within the intergenic vanS(B)-vanY(B) region. Other resistance genes identified were erm(B), mef(E), tet(M), ant(6')-Ia, aph(3')-IIIa and aac(6')-Ie-aph(2'')-Ia. All isolates carried the unexpressed tet(M) gene. The high level of ciprofloxacin resistance was attributable to the first described Gly-61 and Ile-80 mutations in ParC and the Tyr-83 or Arg-83 mutations in GyrA. All isolates contained esp. The presence of hyl was variable. CONCLUSIONS: A large clonal outbreak caused by multidrug-resistant CC17 E. faecium containing pbp5-Tn5382 is described. The persistence of this clone, which has been recovered from both hospital and community settings since 2005, and the possibility of transferring this Tn5382 to other epidemic ampicillin-resistant clonal types currently circulating in Spain might contribute to increasing the prevalence of vancomycin-resistant enterococci in our area. This study constitutes the first description of mef(E) in E. faecium.


Subject(s)
DNA Transposable Elements , Drug Resistance, Multiple, Bacterial , Enterococcus faecium/drug effects , Enterococcus faecium/isolation & purification , Gram-Positive Bacterial Infections/epidemiology , Gram-Positive Bacterial Infections/microbiology , Bacterial Typing Techniques , Carrier State/microbiology , Cluster Analysis , Cross Infection/epidemiology , Cross Infection/microbiology , DNA Fingerprinting , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Enterococcus faecium/classification , Genes, Bacterial , Genotype , Hospitals , Humans , Molecular Sequence Data , Sequence Analysis, DNA , Spain/epidemiology , Virulence Factors/genetics
19.
J Med Microbiol ; 57(Pt 9): 1170-1171, 2008 Sep.
Article in English | MEDLINE | ID: mdl-18719191

ABSTRACT

We describe what we believe to be the first case of biliary sepsis caused by Acinetobacter ursingii. The patient was a healthy woman with no comorbidities who presented with choledocholithiasis and cholangitis. The performance of an endoscopic cholangiopancreatography was the trigger for A. ursingii bacteraemia. This report highlights the inadequacies of conventional phenotypic tests usually available in clinical microbiology laboratories for the identification of Acinetobacter species.


Subject(s)
Acinetobacter Infections/etiology , Bacteremia/etiology , Cholangiopancreatography, Endoscopic Retrograde/adverse effects , Cholangitis/complications , Choledocholithiasis/complications , Acinetobacter/classification , Acinetobacter/isolation & purification , Acinetobacter Infections/drug therapy , Anti-Bacterial Agents/therapeutic use , Bacteremia/drug therapy , Cefotaxime/therapeutic use , Female , Humans , Middle Aged
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