Publications by authors named "Carmen Torres"

437 Publications

Multidrug-resistant Klebsiella pneumoniae harboring extended spectrum β-lactamase encoding genes isolated from human septicemias.

PLoS One 2021 4;16(5):e0250525. Epub 2021 May 4.

Department of Veterinary Sciences, Microbiology and Antibiotic Resistance Team (MicroART), University of Trás-os-Montes and Alto Douro, Vila Real, Portugal.

Klebsiella pneumoniae is a major pathogen implicated in nosocomial infections. Extended-spectrum β-lactamase (ESBL)-producing K. pneumoniae isolates are a public health concern. We aim to characterize the type of β-lactamases and the associated resistance mechanisms in ESBL-producing K. pneumoniae isolates obtained from blood cultures in a Portuguese hospital, as well as to determine the circulating clones. Twenty-two cefotaxime/ceftazidime-resistant (CTX/CAZR) K. pneumoniae isolates were included in the study. Identification was performed by MALDI-TOF MS and the antimicrobial susceptibility testing by disk-diffusion. The screening test for ESBL-production was performed and ESBL-producer isolates were further characterized. The presence of different beta-lactamase genes (blaCTX-M, blaSHV, blaTEM, blaKPC, blaNDM, blaVIM, blaOXA-48, blaCMY-2, blaDHA-1, blaFOX, blaMOX, and blaACC) was analyzed by PCR/sequencing in ESBL-producer isolates, as well as the presence of other resistance genes (aac(6')-Ib-cr, tetA/B, dfrA, qnrA/B/S, sul1/2/3) or integron-related genes (int1/2/3). Multilocus-sequence-typing (MLST) was performed for selected isolates. ESBL activity was detected in 12 of the 22 CTX/CAZR K. pneumoniae isolates and 11 of them carried the blaCTX-M-15 gene (together with blaTEM), and the remaining isolate carried the blaSHV-106 gene. All the blaCTX-M-15 harboring isolates also contained a blaSHV gene (blaSHV-1, blaSHV-11 or blaSHV-27 variants). Both blaSHV-27 and blaSHV-106 genes correspond to ESBL-variants. Two of the CTX-M-15 producing isolates carried a carbapenemase gene (blaKPC2/3 and blaOXA-48) and showed imipenem resistance. The majority of the ESBL-producing isolates carried the int1 gene, as well as sulphonamide-resistance genes (sul2 and/or sul3); the tetA gene was detected in all eight tetracycline-resistant isolates. Three different genetic lineages were found in selected isolates: ST348 (one CTX-M-15/TEM/SHV-27/KPC-2/3-producer isolate), ST11 (two CTX-M-15/TEM/SHV-1- and CTX-M-15-TEM-SHV-11-OXA-48-producer isolates) and ST15 (one SHV-106/TEM-producer isolate). ESBL enzymes of CTX-M-15 or SHV-type are detected among blood K. pneumoniae isolates, in some cases in association with carbapenemases of KPC or OXA-48 type.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0250525PLOS
May 2021

Genomic Analysis of of the Lineage CC130, Including -Carrying MRSA and MSSA Isolates Recovered of Animal, Human, and Environmental Origins.

Front Microbiol 2021 25;12:655994. Epub 2021 Mar 25.

Area of Biochemistry and Molecular Biology, OneHealth-UR Research Group, University of La Rioja, Logroño, Spain.

Most methicillin resistant (MRSA) isolates harboring gene belong to clonal complex CC130. This lineage has traditionally been regarded as animal-associated as it lacks the human specific immune evasion cluster (IEC), and has been recovered from a broad range of animal hosts. Nevertheless, sporadic -MRSA human infections have been reported, with evidence of zoonotic transmission in some cases. The objective of this study was to investigate the whole-genome sequences of 18 CC130 isolates [13 methicillin-resistant (-MRSA) and five methicillin-susceptible (MSSA)] from different sequences types, obtained from a variety of host species and origins (human, livestock, wild birds and mammals, and water), and from different geographic locations, in order to identify characteristic markers and genomic features. Antibiotic resistance genes found among MRSA-CC130 were those associated with the SSCXI element. Most MRSA-CC130 strains carried a similar virulence gene profile. Additionally, six MRSA-CC130 possessed and one MSSA-ST130 had . The MSSA-ST700 strains were most divergent in their resistance and virulence genes. The pan-genome analysis showed that 29 genes were present solely in MRSA-CC130 (associated with SCCXI) and 21 among MSSA-CC130 isolates (associated with phages). The SCCXI, PBP3, GdpP, and AcrB were identical at the amino acid level in all strains, but some differences were found in PBP1, PBP2, PBP4, and YjbH proteins. An examination of the host markers showed that the 3' region of the bacteriophage φ3 was nearly identical to the reference sequence. Truncated gene was also found in -negative strains (two of them carrying -type gene). The gene of wild rabbit isolates included novel mutations. The gene was found in the three MSSA-ST700 strains from small ruminants and in one MSSA-ST130 from a red deer; these strains also carried a -type gene, different from the human and equine variants. Finally, a phylogenetic analysis showed that the three MSSA-ST700 strains and the two MSSA-ST130 strains cluster separately from the remaining MRSA-CC130 strains with the gene as marker for the main lineage. The presence of the human IEC cluster in some -MRSA-CC130 strains suggests that these isolates may have had a human origin.
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http://dx.doi.org/10.3389/fmicb.2021.655994DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8027229PMC
March 2021

Antimicrobial Resistance Genes and Diversity of Clones among ESBL- and Acquired AmpC-Producing Isolated from Fecal Samples of Healthy and Sick Cats in Portugal.

Antibiotics (Basel) 2021 Mar 5;10(3). Epub 2021 Mar 5.

Microbiology and Antibiotic Resistance Team (MicroART), Department of Veterinary Sciences, University of Trás-os-Montes and Alto Douro, 5000-801 Vila Real, Portugal.

The aim of the study was to analyze the mechanisms of resistance in extended-spectrum beta-lactamase (ESBL)- and acquired AmpC (qAmpC)-producing isolates from healthy and sick cats in Portugal. A total of 141 rectal swabs recovered from 98 sick and 43 healthy cats were processed for cefotaxime-resistant (CTX) recovery (in MacConkey agar supplemented with 2 µg/mL cefotaxime). The matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) method was used for identification and antimicrobial susceptibility was performed by a disk diffusion test. The presence of resistance/virulence genes was tested by PCR sequencing. The phylogenetic typing and multilocus sequence typing (MLST) were determined by specific PCR sequencing. CTX isolates were detected in seven sick and six healthy cats (7.1% and 13.9%, respectively). Based on the synergy tests, 11 of 13 CTX isolates (one/sample) were ESBL-producers (ESBL total rate: 7.8%) carrying the following ESBL genes: ( = 3), ( = 3), ( = 2), ( = 2) and ( = 1). Six different sequence types were identified among ESBL-producers (sequence type/associated ESBLs): ST847/CTX-M-9, CTX-M-27, CTX-M-1; ST10/CTX-M-15, CTX-M-27; ST6448/CTX-M-15, CTX-M-55; ST429/CTX-M-15; ST101/CTX-M-1 and ST40/CTX-M-1. Three of the CTX isolates were CMY-2-producers (qAmpC rate: 2.1%); two of them were ESBL-positive and one ESBL-negative. These isolates were typed as ST429 and ST6448 and were obtained in healthy or sick cats. The phylogenetic groups A/B1/D/clade 1 were detected among ESBL- and qAmpC-producing isolates. Cats are carriers of qAmpC (CMY-2)- and ESBL-producing isolates (mostly of variants of CTX-M group 1) of diverse clonal lineages, which might represent a public health problem due to the proximity of cats with humans regarding a One Health perspective.
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http://dx.doi.org/10.3390/antibiotics10030262DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001562PMC
March 2021

Apparent nosocomial adaptation of Enterococcus faecalis predates the modern hospital era.

Nat Commun 2021 03 9;12(1):1523. Epub 2021 Mar 9.

Department of Biostatistics, Faculty of Medicine, University of Oslo, Oslo, Norway.

Enterococcus faecalis is a commensal and nosocomial pathogen, which is also ubiquitous in animals and insects, representing a classical generalist microorganism. Here, we study E. faecalis isolates ranging from the pre-antibiotic era in 1936 up to 2018, covering a large set of host species including wild birds, mammals, healthy humans, and hospitalised patients. We sequence the bacterial genomes using short- and long-read techniques, and identify multiple extant hospital-associated lineages, with last common ancestors dating back as far as the 19th century. We find a population cohesively connected through homologous recombination, a metabolic flexibility despite a small genome size, and a stable large core genome. Our findings indicate that the apparent hospital adaptations found in hospital-associated E. faecalis lineages likely predate the "modern hospital" era, suggesting selection in another niche, and underlining the generalist nature of this nosocomial pathogen.
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http://dx.doi.org/10.1038/s41467-021-21749-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7943827PMC
March 2021

The transcriptional landscape of Shh medulloblastoma.

Nat Commun 2021 03 19;12(1):1749. Epub 2021 Mar 19.

Developmental & Stem Cell Biology Program, The Hospital for Sick Children, Toronto, ON, Canada.

Sonic hedgehog medulloblastoma encompasses a clinically and molecularly diverse group of cancers of the developing central nervous system. Here, we use unbiased sequencing of the transcriptome across a large cohort of 250 tumors to reveal differences among molecular subtypes of the disease, and demonstrate the previously unappreciated importance of non-coding RNA transcripts. We identify alterations within the cAMP dependent pathway (GNAS, PRKAR1A) which converge on GLI2 activity and show that 18% of tumors have a genetic event that directly targets the abundance and/or stability of MYCN. Furthermore, we discover an extensive network of fusions in focally amplified regions encompassing GLI2, and several loss-of-function fusions in tumor suppressor genes PTCH1, SUFU and NCOR1. Molecular convergence on a subset of genes by nucleotide variants, copy number aberrations, and gene fusions highlight the key roles of specific pathways in the pathogenesis of Sonic hedgehog medulloblastoma and open up opportunities for therapeutic intervention.
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http://dx.doi.org/10.1038/s41467-021-21883-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979819PMC
March 2021

Prevalence and Genetic Characteristics of CC398 Isolates From Invasive Infections in Spanish Hospitals, Focusing on the Livestock-Independent CC398-MSSA Clade.

Front Microbiol 2021 9;12:623108. Epub 2021 Feb 9.

Área Bioquímica y Biología Molecular, Universidad de La Rioja, Logroño, Spain.

Background: Livestock-associated (LA)-CC398-MRSA is closely related to pigs, being unfrequently detected in human invasive infections. CC398-MSSA is emerging in human invasive infections in some countries, but genetic and epidemiological characteristics are still scarcely reported.

Objectives: To determine the prevalence of (SA) CC398, both MRSA and MSSA, among blood cultures SA isolates recovered in Spanish hospitals located in regions with different pig-farming densities (PD) and characterize the recovered isolates.

Methods: One thousand twenty-two SA isolates (761 MSSA, 261 MRSA) recovered from blood cultures during 6-12 months in 17 Spanish hospitals (2018-2019) were studied. CC398 lineage identification, detection of -types, and antibiotic resistance, virulence and human immune evasion cluster (IEC) genes were analyzed by PCR/sequencing.

Results: Forty-four CC398-MSSA isolates (4.3% of SA; 5.8% of MSSA) and 10 CC398-MRSA isolates (1% of SA; 3.8% of MRSA) were detected. Eleven -types were found among the CC398-MSSA isolates with t571 and t1451 the most frequent -types detected (75%). Most of CC398-MSSA isolates were Immune-Evasion-Cluster (IEC)-positive (88.6%), tetracycline-susceptible (95.5%) and erythromycin/clindamycin-resistant/(T)-positive (75%). No statistical significance was detected when the CC398-MSSA/MSSA rate was correlated to PD (pigs/km) ( = 0.108). On the contrary, CC398-MRSA isolates were all IEC-negative, predominately -t011 (70%), and the CC398-MRSA/MRSA rate was significantly associated to PD ( < 0.005).

Conclusion: CC398-MSSA is an emerging clade in invasive infections in Spanish hospitals. CC398-MRSA (mostly t011) and CC398-MSSA (mostly t571 and t1451) show important differences, possibly suggesting divergent steps in host-adaptation evolutionary processes. While CC398-MRSA is livestock-associated (lacking IEC-system), CC398-MSSA seems to be mostly livestock-independent, carrying human-adaptation markers.
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http://dx.doi.org/10.3389/fmicb.2021.623108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7945039PMC
February 2021

S. pseudintermedius and S. aureus lineages with transmission ability circulate as causative agents of infections in pets for years.

BMC Vet Res 2021 Jan 21;17(1):42. Epub 2021 Jan 21.

Área Bioquímica y Biología Molecular, Universidad de La Rioja, Logroño, 26006, Spain.

Background: Staphylococcus pseudintermedius (SP) and Staphylococcus aureus (SA) are common colonizers of companion animals, but they are also considered opportunistic pathogens, causing diseases of diverse severity. This study focused on the identification and characterization of 33 coagulase-positive staphylococci isolated from diseased pets (28 dogs and five cats) during 2009-2011 in a veterinary hospital in Spain in order to stablish the circulating lineages and their antimicrobial resistance profile.

Results: Twenty-eight isolates were identified as SP and five as SA. Nine methicillin-resistant (MR) isolates (27%) carrying the mecA gene were detected (eight MRSP and one MRSA). The 55% of SP and SA isolates were multidrug-resistant (MDR). MRSP strains were typed as ST71-agrIII-SCCmecII/III-(PFGE) A (n=5), ST68-agrIV-SCCmecV-B1/B2 (n=2), and ST258-agrII-SCCmecIV-C (n=1). SP isolates showed resistance to the following antimicrobials [percentage of resistant isolates/resistance genes]: penicillin [82/blaZ], oxacillin [29/mecA] erythromycin/clindamycin [43/erm(B)], aminoglycosides [18-46/aacA-aphD, aphA3, aadE], tetracycline [71/tet(M), tet(K)], ciprofloxacin [29], chloramphenicol [29/cat], and trimethoprim-sulfamethoxazole [50/dfrG, dfrK]. The dfrK gene was revealed as part of the radC-integrated Tn559 in two SP isolates. Virulence genes detected among SP isolates were as follow [percentage of isolates]: siet [100], se-int [100], lukS/F-I [100], sec [7], and expB [7]. The single MRSA-mecA detected was typed as t011-ST398/CC398-agrI-SCCmecV and was MDR. The methicillin-susceptible SA isolates were typed as t045-ST5/CC5 (n=2), t10576-ST1660 (n=1), and t005-ST22/CC22 (n=1); the t005-ST22 feline isolate was PVL-positive and the two t045-ST45 isolates were ascribed to Immune Evasion Cluster (IEC) type F. Moreover, the t10576-ST1660 isolate, of potential equine origin, harbored the lukPQ and scneq genes. According to animal clinical history and data records, several strains seem to have been acquired from different sources of the hospital environment, while some SA strains appeared to have a human origin.

Conclusions: The frequent detection of MR and MDR isolates among clinical SP and SA strains with noticeable virulence traits is of veterinary concern, implying limited treatment options available. This is the first description of MRSA-ST398 and MRSP-ST68 in pets in Spain, as well the first report of the dfrK-carrying Tn559 in SP. This evidences that current transmissible lineages with mobilizable resistomes have been circulating as causative agents of infections among pets for years.
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http://dx.doi.org/10.1186/s12917-020-02726-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7819200PMC
January 2021

Antimicrobial resistance in Escherichia coli isolates from frugivorous (Eidolon helvum) and insectivorous (Nycteris hispida) bats in Southeast Nigeria, with detection of CTX-M-15 producing isolates.

Comp Immunol Microbiol Infect Dis 2021 Apr 13;75:101613. Epub 2021 Jan 13.

Area Bioquímica y Biología Molecular, Universidad de La Rioja, Logroño, Spain.

Thirty-five Escherichia coli isolates obtained from the liver, spleen and intestines of 180 frugivorous and insectivorous bats were investigated for antimicrobial resistance phenotypes/genotypes, prevalence of Extended-Spectrum beta-lactamase (ESBL) production, virulence gene detection and molecular typing. Eight (22.9 %) of the isolates were multidrug resistant (MDR). Two isolates were cefotaxime-resistant, ESBL-producers and harbored the bla gene; they belonged to ST10184-D and ST2178-B1 lineages. tet(A) gene was detected in all tetracycline-resistant isolates while int1 (n = 8) and bla (n = 7) genes were also found. Thirty-three of the E. coli isolates were assigned to seven phylogenetic groups, with B1 (45.7 %) being predominant. Three isolates were enteropathogenic E. coli (EPEC) pathovars, containing the eae gene (with the variants gamma and iota), and lacking stx1/stx2 genes. Bats in Nigeria are possible reservoirs of potentially pathogenic MDR E. coli isolates which may be important in the ecology of antimicrobial resistance at the human-livestock-wildlife-environment interfaces. The study reinforces the importance of including wildlife in national antimicrobial resistance monitoring programmes.
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http://dx.doi.org/10.1016/j.cimid.2021.101613DOI Listing
April 2021

Detection of methicillin-resistant coagulase-negative staphylococci and PVL/mecA genes in cefoxitin-susceptible Staphylococcus aureus (t044/ST80) from unpasteurized milk sold in stores in Djelfa, Algeria.

J Dairy Sci 2021 Mar 15;104(3):2684-2692. Epub 2021 Jan 15.

Area Bioquímica y Biología Molecular, Universidad de La Rioja, Madre de Dios 51, 26006 Logroño, Spain. Electronic address:

This study was designed to determine antimicrobial resistance phenotypes and genotypes and virulence factors in Staphylococcus aureus and coagulase-negative staphylococci (CNS) in unpasteurized milk sold in Djelfa, Algeria. Eighty-two unpasteurized cow milk samples were randomly obtained from 82 retail stores in Djelfa and tested to detect staphylococci. Species were identified by biochemical tests and MALDI-TOF. Antimicrobial resistance phenotypes and genotypes were determined by disk diffusion test, PCR, and sequencing. The Staph. aureus isolates were subjected to spa typing, multilocus sequence typing, and detection of virulence genes and the scn gene by PCR and sequencing. Forty-five (54.9%) milk samples were contaminated by staphylococci and 45 isolates were recovered: 10 Staph. aureus (12.2% of total samples) and 35 CNS (42.7%). Resistance to penicillin (blaZ), tetracycline (tetL/tetK), and erythromycin (ermB/msrA/ermC) were the most common phenotypes (genotypes). Three CNS were methicillin-resistant and all were mecA-positive. The Staph. aureus isolates were ascribed to the following lineages [spa type/sequence type/associated clonal complex (number of isolates)]: t267/ST479/CC479 (n = 6), t1510/ST5651/CC45 (n = 1), t359/ST97/CC97/ (n = 1), t346/ST15/CC15 (n = 1), and t044/ST80 (n = 1). The mecA gene was detected in the cefoxitin-susceptible t044/ST80 isolate and co-harbored the lukF/lukS-PV and scn genes. The detection of mecA-PVL-positive Staph. aureus, methicillin-resistant CNS, and multidrug-resistant staphylococcal species indicates a potentially serious health issue and reveals that unpasteurized milk sold in Djelfa city could be a potential vehicle for pathogenic and antimicrobial-resistant staphylococci.
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http://dx.doi.org/10.3168/jds.2020-19270DOI Listing
March 2021

Genetic characterization of extended-spectrum β-lactamase-producing Enterobacteriaceae from a biological industrial wastewater treatment plant in Tunisia with detection of the colistin-resistance mcr-1 gene.

FEMS Microbiol Ecol 2021 03;97(3)

Université de la Manouba, IRESA, École Nationale de Médecine Vétérinaire de Sidi Thabet, Sidi Thabet 2020, Sidi Thabet, Ariana, Tunisia.

This study evaluated the occurrence of extended-spectrum β-lactamases (ESBL) and associated resistance genes, integrons, and plasmid types, as well as the genetic relatedness of enterobacterial isolates in the wastewater treatment plant (WWTP) of La Charguia, Tunis City (Tunisia). A total of 100 water samples were collected at different points of the sewage treatment process during 2017-2019. Antimicrobial susceptibility was conducted by the disc-diffusion method. blaCTX-M, blaTEM and blaSHV genes as well as those encoding non-β-lactam resistance, the plasmid types, occurrence of class1 integrons and phylogenetic groups of Escherichia coli isolates were determined by PCR/sequencing. Genomic relatedness was determined by multi-locus sequence typing (MLST) for selected isolates. In total, 57 ESBL-producer isolates were recovered (47 E. coli, eight Klebsiella pneumoniae, 1 of the Citrobacter freundii complex and 1 of the Enterobacter cloacae complex). The CTX-M-15 enzyme was the most frequently detected ESBL, followed by CTX-M-27, CTX-M-55 and SHV-12. One E. coli isolate harboured the mcr-1 gene. The following phylogroups/sequence types (STs) were identified among ESBL-producing E. coli isolates: B2/ST131 (subclade-C1), A/ST3221, A/ST8900, D/ST69, D/ST2142, D/ST38, B1/ST2460 and B1/ST6448. High numbers of isolates harboured the class 1 integrons with various gene cassette arrays as well as IncP-1 and IncFIB plasmids. Our findings confirm the importance of WWTPs as hotspot collectors of ESBL-producing Enterobacteriaceae with a high likelihood of spread to human and natural environments.
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http://dx.doi.org/10.1093/femsec/fiaa231DOI Listing
March 2021

Human -Carrying MRSA: Clinical Implications and Risk Factors.

Microorganisms 2020 Oct 20;8(10). Epub 2020 Oct 20.

Area of Biochemistry and Molecular Biology, University of La Rioja, 26006 Logroño, Spain.

A new methicillin resistance gene, named , was first described in 2011 in both humans and animals. Since then, this gene has been detected in different production and free-living animals and as an agent causing infections in some humans. The possible impact that these isolates can have in clinical settings remains unknown. The current available information about -carrying methicillin resistant (MRSA) isolates obtained from human samples was analyzed in order to establish its possible clinical implications as well as to determine the infection types associated with this resistance mechanism, the characteristics of these -carrying isolates, their possible relation with animals and the presence of other risk factors. Until now, most human -MRSA infections have been reported in Europe and -MRSA isolates have been identified belonging to a small number of clonal complexes. Although the prevalence of -MRSA human infections is very low and isolates usually contain few resistance (except for beta-lactams) and virulence genes, first isolates harboring important virulence genes or that are resistant to non-beta lactams have already been described. Moreover, severe and even fatal human infection cases have been detected. -carrying MRSA should be taken into consideration in hospital, veterinary and food safety laboratories and in prevention strategies in order to avoid possible emerging health problems.
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http://dx.doi.org/10.3390/microorganisms8101615DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7589452PMC
October 2020

Spp. from Wild Mammals in Aragón (Spain): Antibiotic Resistance Status.

J Vet Res 2020 Sep 16;64(3):373-379. Epub 2020 Sep 16.

Department of Animal Pathology, Infectious Diseases and Epidemiology, Veterinary Faculty of Zaragoza, 50013, Zaragoza, Spain.

Introduction: Antimicrobial resistance is a global health threat. It has been studied in humans and domestic animals, but there is a lack of data on wild animals. The objective of this study is the elucidation of its patterns in spp. isolated from wild mammals of the Autonomous Community of Aragón (Spain).

Material And Methods: A total of 103 mammals (Artiodactyla, Carnivora, Chiroptera, Erinaceomorpha, and Lagomorpha) were studied. A recovery centre provided 32 and hunting 71. Nasal and faecal samples yielded 111 staphylococci, which were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. A susceptibility test to 11 antibiotics was carried out, and statistical analysis was performed.

Results: Some differences were detected in bacterial prevalence depending on how the mammal fed. Artiodactyla, mainly hunted, were predisposed to carry coagulase-positive staphylococci. The staphylococci species recovered were resistant to at least two classes of antibiotics, and were disseminated in all of the geographical areas studied.

Conclusion: Resistant staphylococci are widely distributed in the wild mammals in the areas of the study, but the resistance quantified in them is lower than that to be expected if the use of antibiotics in farms had a direct influence on the wildlife and its environment. On the other hand, resistance to antibiotics restricted to human use was widely disseminated in various wild animal species.
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http://dx.doi.org/10.2478/jvetres-2020-0057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7497752PMC
September 2020

Antibiotic resistance mechanisms in Acinetobacter spp. strains isolated from patients in a paediatric hospital in Mexico.

J Glob Antimicrob Resist 2020 12 8;23:120-129. Epub 2020 Sep 8.

Benemérita Universidad Autónoma de Puebla, Instituto de Ciencias, Posgrado en Microbiología, Centro de Investigaciones de Ciencias Microbiológicas, Puebla, Mexico. Electronic address:

Objectives: The aim of this study was to identify Acinetobacter spp. strains from paediatric patients, to determine their genetic relationship, to detect antibiotic resistance genes and to evaluate the role of efflux pumps in antibiotic resistance.

Methods: A total of 54 non-duplicate, non-consecutive Acinetobacter spp. isolates were collected from paediatric patients. Their genetic relationship, antibiotic resistance profile, efflux pump activity, antibiotic resistance genes and plasmid profile were determined.

Results: The isolates were identified as 24 Acinetobacter haemolyticus, 24 Acinetobacter calcoaceticus-baumannii (Acb) complex and 1 strain each of Acinetobacter junii, Acinetobacter radioresistens, Acinetobacter indicus, Acinetobacter lwoffii, Acinetobacter ursingii and Acinetobacter venetianus. The 24 A. haemolyticus were considered genetically unrelated. One strain was resistant to carbapenems, two to cephalosporins, two to ciprofloxacin and sixteen to aminoglycosides. The antibiotic resistance genes bla (29%), bla (4%), bla (8%), bla (29%), bla (4%), aac(6')-Ig (38%) and the novel variants bla (13%), bla (75%), aac(6')-Iga (4%), aac(6')-Igb (13%) and aac(6')-Igc (42%) were detected. Among 24 Acb complex, 5 were multidrug-resistant, carbapenem-resistant strains carrying bla and bla; they were genetically related and had the same plasmid profile. Other species were susceptible. In some strains of A. haemolyticus and Acb complex, the role of RND efflux pumps was evidenced by a decrease in the MICs for cefotaxime, amikacin and ciprofloxacin in the presence of an efflux pump inhibitor.

Conclusions: This study identified isolates of A. haemolyticus carrying new β-lactamase variants and shows for the first time the contribution of efflux pumps to antibiotic resistance in this species.
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http://dx.doi.org/10.1016/j.jgar.2020.08.014DOI Listing
December 2020

Frequency and Characterization of Antimicrobial Resistance and Virulence Genes of Coagulase-Negative Staphylococci from Wild Birds in Spain. Detection of -Carrying Isolates.

Microorganisms 2020 Aug 29;8(9). Epub 2020 Aug 29.

Área de Bioquímica y Biología Molecular, Universidad de La Rioja, 26006 Logroño, Spain.

The objective of this study was to determine the prevalence and diversity of coagulase-negative staphylococci (CoNS) species from wild birds in Spain, as well as to analyze the antimicrobial resistance phenotype/genotype and the virulence gene content. During 2015-2016, tracheal samples of 242 wild birds were collected in different regions of Spain for staphylococci recovery. The species identification was performed using MALDI-TOF. The antimicrobial resistance phenotype and genotype was investigated by the disk diffusion method and by PCR, respectively. The presence of the virulence genes /-PV, , , , and was investigated by PCR. Moreover, CoNS carrying the gene were subjected to SCC typing. Of the tested animals, 60% were CoNS-carriers, and 173 CoNS isolates were recovered from the 146 positive animals, which belonged to 11 species, with predominance of ( = 118) and ( = 25). A total of 34% of CoNS isolates showed a multidrug resistance phenotype, and 42 -positive methicillin-resistant CoNS (MRCoNS) were detected. The isolates showed resistance to the following antimicrobials (percentage of resistant isolates/antimicrobial resistance genes detected): penicillin (49/ , ), cefoxitin (24/ ), erythromycin and/or clindamycin (92/ (B), (C), (43), (A), (C), (A), (B), (A) and (A)), gentamicin and/or tobramycin (5/ (6')-Ie-(2″)-Ia, (4')-Ia), streptomycin (12/), tetracycline (17/ (K), (L), (M)), ciprofloxacin (4), chloramphenicol (1/ ), fusidic acid (86/ , ) and trimethoprim-sulfamethoxazole (1/ ). None of the isolates harbored the /-PV, , , and genes, but two isolates (1%) carried the gene. Wild birds are frequently colonized by CoNS species, especially . We identified scavenging on intensively produced livestock and feeding on landfills as risk factors for CoNS carriage. High proportions of MRCoNS and multidrug resistant CoNS were detected, which coupled with the presence of important virulence genes is of concern.
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http://dx.doi.org/10.3390/microorganisms8091317DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7564563PMC
August 2020

Genetic characterization of ESBL-producing Escherichia coli and Klebsiella pneumoniae isolated from wastewater and river water in Tunisia: predominance of CTX-M-15 and high genetic diversity.

Environ Sci Pollut Res Int 2020 Dec 7;27(35):44368-44377. Epub 2020 Aug 7.

Departamento de Agricultura y Alimentación, Universidad de La Rioja, Madre de Dios, 53, 26006, Logroño, Spain.

Aquatic environments are crucial hotspots for the dissemination of antibiotic resistant microorganisms and resistance genes. Thus, the purpose of this study was to investigate the occurrence and the genetic characterization of cefotaxime-resistant (CTX) Enterobacteriaceae at a Tunisian semi-industrial pilot plant with biological treatment (WWPP) and its receiving river (Rouriche River, downstream from WWPP) located in Tunis City, during 2017-2018. We collected 105 and 15 water samples from the WWPP and the Rouriche River, respectively. Samples were screened to recover ESBL-producing Enterobacteriaceae (ESBL-E) and isolates were characterized for phenotype/genotype of antimicrobial resistance, integrons, plasmid types and molecular typing (multilocus sequence typing, MLST). Among 120 water samples, 33 and 4 contained ESBL-producing E. coli and K. pneumoniae isolates, respectively. Most isolates were multidrug resistant and produced CTX-M-15 (28 isolates), CTX-M-1 (4 isolates), CTX-M-55 (2 isolates), CTX-M-27 (one isolate), SHV-12 (one isolate) and VEB beta-lactamases (one isolate). All K. pneumoniae were CTX-M-15-positive. Four colistin-resistant isolates were found (MIC 4-8 μg/ml), but they were negative for the mcr genes tested. Class 1 integrons were detected in 21/25 trimethoprim/sulfamethoxazole-resistant isolates, and nine of them carried the gene cassette arrays: aadA2 + dfrA12 (n = 4), aadA1 + dfrA15 (n = 2), aadA5 + dfrA17 (n = 2) and aadA1/2 (n = 1). The IncP and IncFIB plasmids were found in 30 and 16 isolates, respectively. Genetic lineages detected were as follows: E. coli (ST48-ST10 Cplx, ST2499, ST906, ST2973 and ST2142); K. pneumoniae: (ST1540 and ST661). Our findings show a high rate of CTX-M-15 and high genetic diversity of ESBL-E isolates from WWPP and receiving river water.
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http://dx.doi.org/10.1007/s11356-020-10326-wDOI Listing
December 2020

Molecular diversity of Extended-spectrum β-lactamase-producing Escherichia coli from vultures in Canary Islands.

Environ Microbiol Rep 2020 10 31;12(5):540-547. Epub 2020 Aug 31.

Microbiology and Antibiotic Resistance Team (MicroART), Department of Veterinary Sciences, University of Trás-os-Montes and Alto Douro, Vila Real, Portugal.

Antimicrobial resistance among isolates from wild animals is increasingly reported. Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae, and particularly Escherichia coli, have spread worldwide as one of the most common multidrug-resistant organisms. The aim of this study was to determine the carriage rate of ESBL-producing E. coli isolates and their genetic characteristics in wild vultures from the Canary Islands. Faecal samples were collected from 22 apparently healthy free-ranging (wild) vulture chicks from Lanzarote and Fuerteventura (Canary Islands) during July 2019. They were seeded in MacConkey agar supplemented with cefotaxime (2 μg ml ). Colonies with typical morphology of E. coli were identified by MALDI-TOF-MS. Antimicrobial susceptibility was done by disk diffusion. Phenotypic detection of ESBL was performed by double-disk tests. The presence of bla , bla , bla , bla and bla genes, as well as mcr-1 (colistin resistance), tetA/tetB and int1 gene, was tested by PCR/sequencing. Phylogenetic groups and multilocus sequence typing (MLST) were determined by PCR/sequencing. ESBL-producing E. coli isolates were detected in 5/22 tested animals (22.7%), and all isolates (one/animal) carried bla genes: bla (n = 3) and bla (n = 2). ESBL-positive isolates were ascribed to phylogenetic group D (two isolates), B (two isolates) and A (one isolate), and five sequence types were detected (ST/phylogenetic-group/ESBL): ST515/B1/CTX-M-15, ST1290/A/CTX-M-15, ST38/D/CTX-M-15, ST457/D/CTX-M-55 and ST6448/B /CTX-M-55; this suggests a genetic diversity among these isolates. Three CTX-M-15-producing isolates contained the bla gene and one the tetA gene. To our knowledge, this appears to be the first report of ESBL-producing E. coli in vulture chicks from the Canary Islands.
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http://dx.doi.org/10.1111/1758-2229.12873DOI Listing
October 2020

Mechanisms of Linezolid Resistance Among Enterococci of Clinical Origin in Spain-Detection of - and (D)-Carrying .

Microorganisms 2020 Jul 30;8(8). Epub 2020 Jul 30.

Área de Bioquímica y Biología Molecular, Universidad de La Rioja, 26006 Logroño, Spain.

The mechanisms of linezolid resistance among 13 and 6 isolates, recovered from six Spanish hospitals during 2017-2018, were investigated. The presence of acquired linezolid resistance genes and mutations in 23S rDNA and in genes encoding for ribosomal proteins was analyzed by PCR and amplicon sequencing. Moreover, the susceptibility to 18 antimicrobial agents was investigated, and the respective molecular background was elucidated by PCR-amplicon sequencing and whole genome sequencing. The transferability of the linezolid resistance genes was evaluated by filter-mating experiments. The gene was detected in all 13 isolates; and one -positive isolate also carried the recently described (D) gene. Moreover, one isolate displayed the nucleotide mutation G2576T in the 23S rDNA. This mutation was also present in all six isolates. All linezolid-resistant enterococci showed a multiresistance phenotype and harbored several antimicrobial resistance genes, as well as many virulence determinants. The gene was located upstream of the gene in 12 of the isolates. Moreover, an (A)-like gene was located downstream of in two isolates recovered from the same hospital. The gene was transferable in all but one isolates, in all cases along with the gene. The (D) gene was not transferable. The presence of and mutations in the 23S rDNA are the main mechanisms of linezolid resistance among and , respectively. We report the first description of the (D) gene in . The presence of the and (D) genes in Spanish hospitals is a public health concern.
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http://dx.doi.org/10.3390/microorganisms8081155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464793PMC
July 2020

Producing Extended-Spectrum β-lactamases (ESBL) from Domestic Camels in the Canary Islands: A One Health Approach.

Animals (Basel) 2020 Jul 29;10(8). Epub 2020 Jul 29.

Microbiology and Antibiotic Resistance Team (MicroART), Department of Veterinary Sciences, University of Trás-os-Montes and Alto Douro, 5000-801 Vila Real, Portugal.

Objective: This work aimed to determine the carriage rate of ESBL-producing as well as their genetic characteristics in camels from the Canary Islands, Spain.

Methods: Fecal samples were recovered from 58 healthy camels from Gran Canaria (n = 32) and Fuerteventura Islands ( = 26) during July 2019. They were seeded on MacConkey (MC) agar no supplemented and supplemented (MC + CTX) with cefotaxime (2 µg/mL). Antimicrobial susceptibility was determined by disk diffusion test (CLSI, 2018). The presence of , , and genes was tested by PCR/sequencing. Furthermore, the 1 (colistin resistance), A/B (tetracycline resistance), 1 (integrase of class 1 integrons) and genes were analyzed. Phylogenetic groups and sequence types were determined by specific-PCR/sequencing for selected isolates.

Results: was obtained from all the 58 camels in MC media (100%) and in five of them in MC + CTX media (8.6%). Furthermore, 63.8% of isolates recovered from MC agar were susceptible to all the antibiotics tested. The five isolates recovered from MC + CTX media were characterized and two of them were ESBL-producers (3.4%). Both ESBL-producer isolates carried the gene and belonged to the lineages ST3018 (phylogroup A) and ST69 (phylogroup B1). The 3 ESBL-negative isolates recovered from MC-CTX plates were ascribed to phylogroup-B.

Conclusions: Camels can be a source of ESBL-producer bacteria, containing the widespread gene associated with the lineages ST3018 and ST69.
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http://dx.doi.org/10.3390/ani10081295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7459641PMC
July 2020

Effects of partial reinforcement on autoshaping in inbred Roman high- and low-avoidance rats.

Physiol Behav 2020 10 30;225:113111. Epub 2020 Jul 30.

Department of Biopsychology, Ruhr-Universität Bochum, 44801, Bochum, Germany. Electronic address:

Individuals trained under partial reinforcement (PR) typically show a greater resistance to extinction than individuals exposed to continuous reinforcement (CR). This phenomenon is referred to as the PR extinction effect (PREE) and is interpreted as a consequence of uncertainty-induced frustration counterconditioning. In this study, we assessed the effects of PR and CR in acquisition and extinction in two strains of rats, the inbred Roman high- and low-avoidance (RHA and RLA, respectively) rats. These two strains mainly differ in the expression of anxiety, the RLA rats showing more anxiety-related behaviors (hence, more sensitive to frustration) than the RHA rats. At a neurobiological level, mild stress is known to elevate corticosterone in RLA rats and dopamine in RHA rats. We tested four groups of rats (RHA/CR, RHA/PR, RLA/CR, and RLA/PR) in two successive acquisition-extinction phases to try to consolidate the behavioral effects. Animals received training in a Pavlovian autoshaping procedure with retractable levers as the conditioned stimulus, food pellets as the unconditioned stimulus, and lever presses as the conditioned response. In Phase 1, we observed a PREE in lever pressing in both strains, but this effect was larger and longer lasting in RHA/PR than in RLA/PR rats. In Phase 2, reacquisition was fast and the PREE persisted in both strains, although the two PR groups no longer differed in lever pressing. The results are discussed in terms of frustration theory and of uncertainty-induced sensitization of dopaminergic neurons.
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http://dx.doi.org/10.1016/j.physbeh.2020.113111DOI Listing
October 2020

Diagnostic Accuracy of Interferon Gamma-Induced Protein 10 mRNA Release Assay for Tuberculosis.

J Clin Microbiol 2020 09 22;58(10). Epub 2020 Sep 22.

Department of Infectious Disease Immunology, Centre for Vaccine Research, Statens Serum Institut, Copenhagen, Denmark

Interferon gamma (IFN-γ) release assays (IGRAs) are increasingly used to test for latent tuberculosis (TB) infection. Although highly specific, IGRAs have a relatively high false-negative rate in active TB patients. A more sensitive assay is needed. IFN-γ-induced protein 10 (IP-10) is an alternative biomarker with a 100-fold-higher expression level than IFN-γ, allowing for different analysis platforms, including molecular detection. The PCR technique is already an integrated tool in most TB laboratories and, thus, an obvious platform to turn to. In this case-control study, we investigated the diagnostic sensitivity and specificity of a molecular assay detecting IP-10 mRNA expression following antigen stimulation of a blood sample. We included 89 TB patients and 99 healthy controls. Blood was drawn in QuantiFeron-TB gold in-tube (QFT) assay tubes. Eight hours poststimulation, IP-10 mRNA expression was analyzed, and 20 h poststimulation, IP-10 and IFN-γ protein plasma levels were analyzed using an in-house IP-10 enzyme-linked immunosorbent assay (ELISA) and the official QFT ELISA, respectively. The IP-10 mRNA assay provided high specificity (98%), sensitivity (80%), and area under the concentration-time curve (AUC) (0.97); however, the QFT assay provided a higher overall diagnostic potential, with specificity of 100%, sensitivity of 90%, and AUC of 0.99. The IP-10 protein assay performed on par with the QFT assay, with specificity of 98%, sensitivity of 87%, and AUC of 0.98. We have provided proof of high technical performance of a molecular assay detecting IP-10 mRNA expression. As a diagnostic tool, this assay would gain from further optimization, especially on the kinetics of IP-10 mRNA expression.
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http://dx.doi.org/10.1128/JCM.00848-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7512164PMC
September 2020

Antimicrobial resistance and virulence of Pseudomonas spp. among healthy animals: concern about exolysin ExlA detection.

Sci Rep 2020 07 15;10(1):11667. Epub 2020 Jul 15.

Área de Microbiología Molecular, Centro de Investigación Biomédica de La Rioja (CIBIR), C/Piqueras 98, 3ªplanta, 26006, Logroño, Spain.

Pseudomonas is a ubiquitous genus that also causes human, animal and plant diseases. Most studies have focused on clinical P. aeruginosa strains from humans, but they are scarce on animal strains. This study was aimed to determine the occurrence of Pseudomonas spp. among faecal samples of healthy animals, and to analyse their antimicrobial resistance, and pathogenicity. Among 704 animal faecal samples analysed, 133 Pseudomonas spp. isolates (23 species) were recovered from 46 samples (6.5%), and classified in 75 different PFGE patterns. Low antimicrobial resistance levels were found, being the highest to aztreonam (50.3%). Five sequence-types (ST1648, ST1711, ST2096, ST2194, ST2252), two serotypes (O:3, O:6), and three virulotypes (analysing 15 virulence and quorum-sensing genes) were observed among the 9 P. aeruginosa strains. Type-3-Secretion System genes were absent in the six O:3-serotype strains that additionally showed high cytotoxicity and produced higher biofilm biomass, phenazine pigments and motility than PAO1 control strain. In these six strains, the exlAB locus, and other virulence genotypes (e.g. RGP69 pathogenicity island) exclusive of PA7 outliers were detected by whole genome sequencing. This is the first description of the presence of the ExlA exolysin in P. aeruginosa from healthy animals, highlighting their pathological importance.
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http://dx.doi.org/10.1038/s41598-020-68575-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7363818PMC
July 2020

Extended Spectrum β-Lactamase-Producing and from Broiler Liver in the Center of Algeria, with Detection of CTX-M-55 and B2/ST131-CTX-M-15 in .

Microb Drug Resist 2021 Feb 29;27(2):268-276. Epub 2020 Jun 29.

Area Bioquímica y Biología Molecular, Universidad de La Rioja, Logroño, Spain.

This study aimed to determine the prevalence and diversity of extended-spectrum β-lactamase (ESBL)-producing and multidrug-resistant (MDR) and isolates from 136 broiler livers randomly purchased in 136 retail markets in Djelfa (Algeria). Isolation was performed on Hektoen agar and bacterial identification was carried out by API20E system and Maldi-TOF-MS (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry). Antimicrobial susceptibility was tested by the disk diffusion and agar dilution methods. Detection of ESBLs and other resistance and integron genes, phylogenetic grouping, and molecular typing was performed by PCR and sequencing. Seventy-eight isolates (one per positive sample) were recovered: 73 and 5 . Among , 86.3% of isolates were MDR. ESBL activity was revealed in eight and five e isolates (rates of 5.9% and 3.7% in analyzed samples, respectively). ESBL genes detected among were as follows (number of isolates): (3), (3), (1), and (1); all ESBL-producing isolates carried the gene. ESBL-producing isolates were assigned to lineages (phylogroup/sequence type and number of isolates in parenthesis): A/ST48 (1), B1/ST6448 (1), B1/ST5087 (3), B1/ST23 (1), and B2/ST131 (two isolates). isolates were ascribed to sequence types ST2010 and ST3483. Regarding the 65 non-ESBL isolates, the most observed resistance genes were as follows: (A) (75%), (57.1%), and (43.5%). Class1 integrons were revealed in seven non-ESBL isolates (10.7%) and two gene-cassette arrays were identified: 1 and 11. Our study provides evidence that broiler-derived food from Center of Algeria constitutes a source of ESBL and/or MDR-producing Enterobacteriaceae, with detection of relevant ESBL genes and epidemic clones.
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http://dx.doi.org/10.1089/mdr.2020.0024DOI Listing
February 2021

Staphylococci among Wild European Rabbits from the Azores: A Potential Zoonotic Issue?

J Food Prot 2020 Jul;83(7):1110-1114

Microbiology and Antibiotic Resistance Team (MicroART), Department of Veterinary Sciences, University of Trás-os-Montes and Alto Douro, Quinta de Prados 500-801, Vila Real, Portugal.

Abstract: The prevalence and diversity of Staphylococcus species from wild European rabbits (Oryctolagus cuniculus) in the Azores were investigated, and the antibiotic resistance phenotype and genotype of the isolates were determined. Nasal samples from 77 wild European rabbits from São Jorge and São Miguel islands in Azores were examined. Antibiotic susceptibility of the isolates was determined using the Kirby-Bauer disk diffusion method, and the presence of antimicrobial resistance genes and virulence factors was determined by PCR. The genetic lineages of S. aureus isolates were characterized by spa typing and multilocus sequence typing. A total of 49 staphylococci were obtained from 35 of the 77 wild rabbits. Both coagulase-positive (8.2%) and coagulase-negative (91.8%) staphylococci were detected: 4 S. aureus, 17 S. fleurettii, 13 S. sciuri, 7 S. xylosus, 4 S. epidermidis, and 1 each of S. simulans, S. saprophyticus, S. succinus, and S. equorum. The four S. aureus isolates showed methicillin susceptibility and were characterized as spa type t272/CC121, Panton-Valentine leukocidin negative, and hlB positive. Most of the coagulase-negative staphylococci showed resistance to fusidic acid and beta-lactams, and multidrug resistance was identified especially among S. epidermidis isolates. The mecA gene was detected in 20 isolates of the species S. fleurettii and S. epidermidis, associated with the blaZ gene in one S. epidermidis isolate. Five antimicrobial resistance genes were detected in one S. epidermidis isolate (mecA,dfrA,dfrG,aac6'-aph2'', and ant4). Our results highlight that wild rabbits are reservoirs or "temporary hosts" of Staphylococcus species with zoonotic potential, some of them carrying relevant antimicrobial resistances.

Highlights:
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http://dx.doi.org/10.4315/0362-028X.JFP-19-423DOI Listing
July 2020

Mechanisms of Linezolid Resistance Among Clinical spp. in Spain: Spread of Methicillin- and Linezolid-Resistant ST2.

Microb Drug Resist 2021 Feb 22;27(2):145-153. Epub 2020 May 22.

Área de Bioquímica y Biología Molecular, Universidad de La Rioja, Logroño, Spain.

This study aimed at determining the mechanisms of linezolid resistance and the molecular characteristics of clinical ( = 2) and coagulase-negative staphylococci ( = 15) isolates obtained from four Spanish hospitals. The detection of linezolid resistance mechanisms (mutations and acquisition of resistance genes) was performed by PCR/sequencing. The antimicrobial resistance and virulence profile was determined, and the isolates were typed by different molecular techniques. Moreover, the genetic environment of the gene was determined by whole-genome sequencing. The gene was detected in one methicillin-resistant (MRSA) that also displayed the amino acid change Val118Ala in the ribosomal protein L4. The second isolate was methicillin susceptible and showed different alterations in the ribosomal protein L4. All remaining linezolid-resistant ( = 14) and isolates ( = 1) showed the mutation G2576T ( = 14) or C2534T ( = 1) in the 23S rRNA. Moreover, different amino acid changes were detected in the ribosomal proteins L3 and L4 in isolates. All isolates belonged to the multilocus sequence type ST2. Linezolid-resistant staphylococci (LRS) showed a multiresistance phenotype, including methicillin resistance that was detected in all isolates but one, and was mediated by the gene. The gene in the MRSA isolate was located together with the gene on a conjugative 38,864 bp plasmid. Linezolid- and methicillin-resistant ST2 showing mutations in the 23S rRNA and in the ribosomal proteins L3 and L4 are spread among Spanish hospitals, whereas LRS carrying acquired linezolid resistance genes are rarely detected.
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http://dx.doi.org/10.1089/mdr.2020.0122DOI Listing
February 2021

Simultaneous Nasal Carriage by Methicillin-Resistant and Methicillin Susceptible of Lineage ST398 in a Live Pig Transporter.

Pathogens 2020 May 21;9(5). Epub 2020 May 21.

Department of Veterinary Medicine, University of Cambridge, Cambridge CB3 0ES, UK.

Methicillin-resistant (MRSA) sequence type (ST)398 is a livestock associated (LA) lineage with zoonotic potential, especially in humans with live pig contact. The objective of this study was to characterize two strains of lineage ST398 (one methicillin-resistant (MRSA), one methicillin-susceptible (MSSA)) isolated from the same nasal sample of a patient admitted in the Intensive-Care Unit of a Spanish Hospital, and with previous occupational exposure to live pigs, by whole-genome-sequencing (WGS). The sample was obtained during routine surveillance for MRSA colonization. Purified genomic DNA was sequenced using Illumina HiSeq 2000 and processed using conventional bioinformatics software. The two isolates recovered were both t011/ST398 and showed similar resistance-phenotypes, other than methicillin susceptibility. The possession of antibiotic resistance genes was the same, except for the A-gene located in SCCV in the MRSA isolate. The MSSA isolate harbored remnants of a SCC following the deletion of 17342bp from a recombination between two putative primases. Both isolates belonged to the livestock-associated clade as defined by three canonical single-nucleotide-polymorphisms, and neither possessed the human immune evasion cluster genes, , or The core genome alignment showed a similarity of 99.6%, and both isolates harbored the same mobile genetic elements. The two nasal ST398 isolates recovered from the patient with previous occupational exposure to pigs appeared to have a livestock origin and could represent different evolutionary steps of animal-human interface lineage. The MSSA strain was formed as a result of the loss of the A gene from the livestock-associated-MRSA lineage.
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http://dx.doi.org/10.3390/pathogens9050401DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7281718PMC
May 2020

Effects of alcohol consumption induced by reward loss on behavior in the hole-board test.

Behav Processes 2020 Jul 11;176:104135. Epub 2020 May 11.

Department of Psychology, University of Jaén, Jaén 23071, Spain. Electronic address:

Rats exposed to reward downshift (from 32 to 4% sucrose) increase 2% alcohol intake in a 2-h, free-choice preference test which also offered water. This effect was accompanied by augmented general activity in the elevated plus maze (Donaire et al., 2018, Behav Proc, 150, 59-65). In the present study we analyzed the effect of alcohol consumption induced by reward downshift on anxiety behaviors registered in the hole-board (HB) test. Sixteen food-deprived female Wistar rats received 32% sucrose for ten 5-min daily sessions and were then downshifted to 4% sucrose for two 5-min daily sessions (postshift). Sessions also involved testing animals in a 2-h, 2-bottle preference task with 2% alcohol vs. water (Group A), or water vs. water (Group W). On postshift sessions, animals were exposed to a 6-min HB test after the preference task. Reward devaluation significantly reduced sucrose intake in Groups A and W, and increased alcohol consumption in Group A, but had no effect on water consumption in Group W. Increased alcohol consumption was followed by higher head-dipping frequency in the HB test compared with Group W. The results are discussed in terms of the impact of reward loss on anxiety behaviors in the HB test and the anxiolytic effects of alcohol in situations involving negative affect.
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http://dx.doi.org/10.1016/j.beproc.2020.104135DOI Listing
July 2020

Detection of a cfr-positive MRSA CC398 strain in a pig farmer in Spain.

Enferm Infecc Microbiol Clin 2021 Mar 6;39(3):139-141. Epub 2020 May 6.

Área de Bioquímica y Biología Molecular, Universidad de La Rioja, Logroño, Spain. Electronic address:

Objective: To characterize one linezolid- and methicillin-resistant Staphylococcus aureus (MRSA) isolate recovered from a nasal sample of a pig farmer patient.

Methods: The detection of linezolid resistance mechanisms was performed by PCR and sequencing. The antimicrobial resistance and virulence profile was investigated, and the molecular typing was performed by molecular techniques. The transference of cfr gene was assessed by conjugation experiments and its genetic environment was investigated by specific PCRs.

Results: The linezolid-resistant MRSA isolate was typed as t011-ST398/CC398-SCCmecV-agrI and carried the cfr gene. The isolate was multidrug-resistant but lacked the virulence genes studied. The cfr gene was co-located with the fexA gene on a Tn558 variant and was successfully transferred by conjugation.

Conclusion: We report the first description of LA-MRSA-CC398 carrying the cfr gene in Spain. This finding highlights the importance of surveillance programmes to determine the presence and spread of the cfr gene in the livestock and clinical settings.
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http://dx.doi.org/10.1016/j.eimc.2020.03.006DOI Listing
March 2021

Occurrence of Organochlorine Pesticides in Human Tissues Assessed Using a Microextraction Procedure and Gas Chromatography-Mass Spectrometry.

J Anal Toxicol 2021 Feb;45(1):84-92

Department of Analytical Chemistry, Faculty of Chemistry, Regional Campus of International Excellence "Campus Mare Nostrum", University of Murcia, E-30100 Murcia, Spain.

This work focuses on the development, validation and application of an analytical method for the determination of twenty organochlorine pesticides (OCPs) in human tissues using salting-out liquid-liquid extraction and dispersive liquid-liquid microextraction for sample preparation and gas chromatography-mass spectrometry to analyze the obtained extracts. Measurement of the concentration levels of these toxics in tissues can be used to assess the risk of the population to exposure. The linearity of the proposed method was verified in the 10-1,000 ng/g range. The sensitivity was evaluated calculating the limits of detection (LODs) for 20 OCPs (α-, β-, γ- and δ-hexachlorocyclohexane (HCH), α- and β-endosulfan, endosulfan sulfate, aldrin, dieldrin, endrin, endrin ketone, endrin aldehyde, α- and γ-chlordane, 4,4'-dichlorodiphenyltrichloroethane, 4,4'-dichlorodiphenyldichloroethylene (DDE), 4,4'-dichlorodiphenyldichloroethane, heptachlor, heptachlor epoxide and methoxychlor), most of them being found between 1.0 and 16 ng/g. The intra- and interday precisions were <12% for relative standard deviation values. The accuracy of the method was evaluated by recovery studies, which gave recovery percentages in the 85-109% range. Seven different tissues (liver, kidney, heart, spleen, lung, brain and abdominal fat) from eight autopsies were analyzed, and only three cases were seen to have β-HCH and 4,4'-DDE in abdominal fat, while 4,4'-DDE was also detected in the heart of one case. The rest of the samples were free of the studied OCPs at least above the corresponding LODs.
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http://dx.doi.org/10.1093/jat/bkaa036DOI Listing
February 2021

Coagulase-negative staphylococci carrying cfr and PVL genes, and MRSA/MSSA-CC398 in the swine farm environment.

Vet Microbiol 2020 Apr 28;243:108631. Epub 2020 Feb 28.

Department of Food and Agriculture, University of La Rioja, Logroño, Spain. Electronic address:

This work aimed at characterizing four Staphylococcus aureus and 68 coagulase-negative staphylococci (CoNS), recovered from the air and liquid manure tank of two swine farms with intensive- and semi-extensive-production types, for their antimicrobial resistance pheno-/genotypes and their virulence gene content. Molecular typing was performed by spa typing, MLST, agr typing, and SCCmec typing, where applicable. Conjugation experiments were performed to assess the transferability of the linezolid resistance gene cfr, and its genetic environment was determined by Whole-Genome-Sequencing. The four S. aureus (intensive-production farm, IP-farm) were typed as t011-agrI-CC398-ST398, were scn-negative and two of them were methicillin-resistant (MRSA) with the mecA gene (SCCmec-V). Multidrug resistance was seen in 87 % of the CoNS. Statistically significant differences among the antimicrobial resistance rates of CoNS from the two farms were observed for cefoxitin, aminoglycosides, tetracycline, ciprofloxacin and trimethoprim-sulfamethoxazole. Eight methicillin-resistant CoNS, which were recovered from the IP-farm, carried the mecA gene. One S. simulans isolate was PVL-positive and three S. cohnii eta-positive. One S. equorum and one S. arlettae showed linezolid resistance and carried the cfr gene (IP-farm), which was non-transferable by conjugation into S. aureus. The cfr genetic context in both isolates was identical, with the lsa(B) gene located upstream of cfr. The environment of swine farms might contribute to the dissemination of CoNS that show multidrug resistance and harbor important virulence factors.
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http://dx.doi.org/10.1016/j.vetmic.2020.108631DOI Listing
April 2020