Cristina Seral

Correlation between the presence of symptoms and the Giardia duodenalis genotype

Clinical manifestations of Giardia duodenalis infection vary from asymptomatic infection to chronic diarrhoea. We study the correlation between the presence of symptoms and the G. duodenalis genotype in 108 patients with giardiasis. Patient age ranged from 2 to 72xa0years old. We found a correlation between assemblage AII and symptomatic infections, and between assemblage B and asymptomatic infections in the overall patient group and in patients less than five years of age. Nevertheless, if only patients of more than five years of age were considered, no statistically significant relationship between assemblage and symptomatic or asymptomatic Giardia infections was found. In these patients, host factors may affect the presence of clinical manifestations more than Giardia assemblage.

Evaluation of an immunochromatographic dip strip test for simultaneous detection of Cryptosporidium spp , Giardia duodenalis , and Entamoeba histolytica antigens in human faecal samples

Immunochromatographic (IC) tests may play an important role in the future diagnosis of parasitic diseases because of their speed and simplicity of use. A recently developed test to detect Cryptosporidium spp, Giardia duodenalis and Entamoeba histolytica was evaluated. Microscopy and PCR were the “gold standard” reference techniques and the results of this IC test were compared with those obtained with ELISA and IC single test for the three parasites. One hundred sixty stool samples were assayed. Using microscopy, 22 samples were diagnosed as positive for Cryptosporidium spp., 31 for Giardia duodenalis, 41 for Entamoeba histolytica/dispar, and 68 had a negative diagnosis for the three parasites. Results of IC tests show sensitivities of 70–72% for Cryptosporidium, 90–97% for Giardia and 62.5% for Entamoeba histolytica. Specificities were of 93.6–94.9%, >99% and 96.1%, respectively. In all diagnoses, agreement with microscopy and PCR was over 90%, except in the triple test and microscopy in E. histolytica detection that was 76.3%, due to the inability of microscopy to differentiate E. histolytica from nonpathogenic species such as E. dispar or E. moshkovskii. The triple stool immunoassays provide adequate sensitivities and specificities for use in outbreak situations, for screening proposals and for massive assays in endemic areas where a large number of samples must be analysed or as complementary test for individual diagnosis.

Nosocomial outbreak of methicillin- and linezolid-resistant Staphylococcus epidermidis associated with catheter-related infections in intensive care unit patients

A total of 128 isolates associated with catheter-related infections was recovered from 101 intensive care unit patients in a Spanish hospital during March 2008 to August 2009, and 27 of these isolates (from 21 patients) were typed as methicillin- and linezolid-resistant Staphylococcus epidermidis. Thirteen of these 21 patients (62%) had received linezolid during the 3 months preceding S. epidermidis recovery. Two closely related pulsotypes (P1a and P1b) were identified among the 27 studied isolates that belonged to the sequence type ST2 and harboured the mecA gene and the SCCmecIII type. The strains recovered from patients 1-4 (pulsotype P1a) showed the nucleotide mutation G2474T inside the amplified fragment of the 23S rRNA region and carried the aac(6)-Ie-aph(2″)-Ia, ant(4), and catA genes, whereas the strains from patients 5-21 (pulsotype P1b) showed the mutation G2603T and carried the aac(6)-Ie-aph(2″)-Ia gene. None of the strains amplified the cfr gene. The ica gene and the IS256 element were detected in all strains. The emergence of 2 closely related methicillin- and linezolid-resistant S. epidermidis strains with 2 different mutations in the 23S rRNA region (G2474T and G2603T) is reported in this study as a cause of a nosocomial outbreak. The presence of G2474T or G2603T point mutations suggests that there are multiple potential sites within domain V of the 23S rRNA region at which mutations could confer resistance to linezolid in the clinical isolates. To our knowledge, this is the first report in which the G2474T mutation has been detected in domain V of the 23S rRNA gene of clinical S. epidermidis.

Carbapenem-resistant Pseudomonas aeruginosa strains from a Spanish hospital: Characterization of metallo-beta-lactamases, porin OprD and integrons

Molecular typing and mechanisms of carbapenem resistance such as alterations in porin OprD and presence of metallo-beta-lactamases (MBLs), as well as integrons have been studied in a collection of carbapenem-resistant Pseudomonas aeruginosa (CRPA) isolates from a Spanish hospital. One hundred and twenty-three CRPA isolates were recovered from different samples of 80 patients. Clonal relationship among CRPA was analyzed by SpeI-PFGE. Susceptibility testing to 11 antibiotics and MBL phenotype was determined by microdilution, IP/IPI E-test and double disc method. The oprD gene was studied by PCR and sequencing, and mutations were determined comparing with P. aeruginosa PAO1 sequence. Characterization of MBLs, and class 1 and 2 integrons were studied by PCR and sequencing. SDS-PAGE analysis of outer membrane proteins of selected strains was performed. Seventy-four-per-cent of patients with CRPA were hospitalised in the ICU setting and 50% had long hospitalization stays. Sixty-four different PFGE patterns were detected, and 87 CRPA strains were further analyzed. MBL phenotype was detected in 43 of 87 strains (49.4%), which contained blaVIM-2 gene inside class 1 integrons. VIM-2-producing strains belonged to lineages ST175, ST235, and ST973. A great diversity of nucleotide insertions, deletions, and mutations in oprD gene, and the presence of a new insertion sequence (ISPa45) truncating oprD were identified among CRPA strains. Class 1 integrons were detected in 75% of CRPA strains, blaVIM-2 and the new arrangement aac(3)-Ia+ISPa34+aadA1 (named as In661) being the most frequent gene-cassette arrays detected. Other gene cassettes detected in integrons were: aadB, aadA6, aadA7, aac(6)-Ib, and blaOXA-46.

Detection and characterization of a ST6 clone of vanB2-Enterococcus faecalis from three different hospitals in Spain.

Thirteen vancomycin-resistant and teicoplanin-susceptible Enterococcus faecalis isolates were recovered from unrelated patients in three Spanish hospitals from November 2009 to December 2010. All isolates carried the vanB2 gene, showed indistinguishable or closely-related PFGE patterns and were ascribed to the sequence type ST6 (included into the high-risk clonal-complex CC2). They showed a multiresistance phenotype (erythromycin, tetracycline, ciprofloxacin and high-level-resistance to streptomycin, gentamicin and kanamycin) and harboured the aac(6’)-aph(2”), ant(6)-Ia, and tet(M)+/−tet(L) genes. All isolates produced gelatinase and harboured the gelE gene, but not the esp or hyl genes. The inclusion of the vanB2 gene into the Tn5382 transposon was demonstrated in one isolate. Clonal dissemination of vanB2-containing the E. faecalis strain is demonstrated.

Epidemiological features, resistance genes, and clones among community-onset methicillin-resistant Staphylococcus aureus (CO-MRSA) isolates detected in northern Spain

Twenty-nine community-onset methicillin-resistant Staphylococcus aureus (CO-MRSA) isolates were prospectively selected according to epidemiological criteria among 374 MRSA isolates collected in our laboratory during 2009-2010 in order to determine which community-associated MRSA (CA-MRSA) and healthcare-associated MRSA (HA-MRSA) clones are circulating in the community in northern Spain. PVL genes were detected in 5 strains (17.2%) that belonged to SCCmec type IV or V and to the agr group I (ST8 and ST2050), agr group II (ST121), and agr group III (ST30 and ST852). These strains were isolated from patients with different clinical manifestations such as urinary tract infection, abscess, or pneumonia, and most of them belonged to emergency department patients with no history of visits to General Practitioners (GPs) in the year before the isolation. We considered that the prevalence of CA-MRSA in community-onset isolates was low (17.2%). A high proportion of the CO-MRSA strains (58.6%) were ST125-MRSA-IVc (CC5), responsible for most of the infections caused by HA-MRSA strains in Spain. This endemic clone is also circulating in the community of northern Spain as we could demonstrate in this study. Antimicrobial resistance was found in spa type t067 isolates linked to the presence of ant(4)-Ia and msr(A). Most of the CO-MRSA isolates in this study corresponded to spa types more associated to the hospital environment, suggesting the interchange of genetic lineages of MRSA among community and hospital niches.

Molecular mechanisms of quinolone resistance in clinical isolates of Aeromonas caviae and Aeromonas veronii bv. sobria.

Mutations in quinolone targets were studied together with quinolone efflux pump activation and plasmid-mediated quinolone resistance determinants in nalidixic-acid-resistant isolates of Aeromonas caviae and Aeromonas veronii. Among 135 clinical Aeromonas spp. isolated from stools of patients with gastrointestinal symptoms, 40 nalidixic acid-resistant strains belonging to A. caviae and A. veronii were selected and their susceptibility to different quinolones (ciprofloxacin, norfloxacin, ofloxacin) further evaluated. Susceptibility to nalidixic acid and ciprofloxacin in the presence/absence of Phe- Arg-β-naphthylamide was also determined. The 16 nalidixic-acid-resistant strains identified as A. caviae were more resistant than the 24 A. veronii bv. sobria strains to ciprofloxacin, norfloxacin, and ofloxacin. All strains showed a mutation (single or double) at position 83 of the QRDR sequence of gyrA, with Ser-83 → Ile as the most frequent substitution. By contrast, no mutations were found at position 87 of gyrA. Double substitutions (GyrA-ParC) were detected in 50% of A. veronii bv. sobria isolates and in 43.75% of A. caviae strains. Both species showed decreases in the MICs of ciprofloxacin. A qnrS gene was found in an A. caviae strain. Thus, in the two species of nalidixic-acid-resistant Aeromonas isolates examined, resistance mediated by efflux pumps contributed only slightly to ciprofloxacin resistance. While two isolates were positive for the aac(6)-Ib gene, no -cr variants were detected.

Prevalencia de Giardia duodenalis genotipo B en humanos en Zaragoza y León, España

OBJECTIVESnTo study Giardia duodenalis assemblages circulating in the health areas of two hospitals in Zaragoza and León (Spain).nnnMETHODSnA total of 211 stool samples with Giardia were genotyped by PCR of the tpi gene.nnnRESULTSnAssemblage B was the most prevalent, both in Zaragoza (84,7%) and León (95,1%). The remaining isolates were identified as AII+B.nnnCONCLUSIONSnWe detected the spread of G. duodenalis assemblage B in Zaragoza and in León, with an increase in its prevalence in Zaragoza compared to previous studies.

Emerging and reemerging pathogens.

From 1973 to 1995, 29 new and reemerging pathogenic microbes were recognized. However, in discussions about emerging infectious diseases, the focus is often on the clinical effects of the host-parasite relationship, rather than the examination of the biology of the pathogen. Many of what we refer to as emerging diseases are characterized better as diseases of human progress. Thus, the aerosolization of water has played an important role in the emergence of Legionella pneumophila infections. New diseases are superimposed on endemic diseases such as diarrhoeal diseases, malaria and tuberculosis. In addition, many pathogens are becoming increasingly resistant to standard antimicrobial drugs, making treatment difficult and in some cases impossible. We summarize our experience on emerging parasitic diseases (primary amoebic meningoencephalitis, respiratory cryptosporidiosis, and diplogonoporiasis), and selected problems of bacterial resistance (MDR tuberculosis caused by Mycobacterium bovis and macrolide-resistance mechanisms of Streptococcus pneumoniae and S. pyogenes).

Comparison of Local Features from Two Spanish Hospitals Reveals Common and Specific Traits at Multiple Levels of the Molecular Epidemiology of Metallo-β-Lactamase-Producing Pseudomonas spp.

ABSTRACT Twenty-seven well-characterized metallo-β-lactamase (MBL)-producing Pseudomonas strains from two distantly located hospitals were analyzed. The results revealed specific features defining the multilevel epidemiology of strains from each hospital in terms of species, clonality, predominance of high-risk clones, composition/diversity of integrons, and linkages of Tn402-related structures. Therefore, despite the global trends driving the epidemiology of MBL-producing Pseudomonas spp., the presence of local features has to be considered in order to understand this threat and implement proper control strategies.