Tracking methicillin-resistant Staphylococcus aureus clones during a 5-year period (1998 to 2002) in a Spanish hospital

Three hundred seventy-five consecutive methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates recovered between 1998 and 2002 at the Nuestra Señora de Candelaria University Hospital in Tenerife, Spain, were analyzed by molecular fingerprinting techniques to determine MRSA clonal types and their prevalence over time. After determining antibiotic susceptibility, we used SmaI-digested genomic DNA separated by pulsed-field gel electrophoresis (PFGE) to characterize MRSA isolates and to establish PFGE types. Additionally, several selected isolates representative of each major PFGE type were tested by multilocus sequence typing (MLST) and by a multiplex PCR method capable of identifying the structural type of the staphylococcal cassette chromosome mec (SCCmec), generating the corresponding sequence type (ST)-SCCmec types. Results of PFGE, supported by those of MLST and SCCmec typing, allowed us to identify six MRSA clones within the five major PFGE types and document temporal shifts in the prevalence of these MRSA clones from 1998 to 2002. Four of the clones were the pandemic “Iberian” (designated ST247-MRSA-IA), EMRSA-15 (ST22-MRSA-IV), EMRSA-16 (ST36-MRSA-II), and the so-called pediatric (ST5-MRSA-IV) clones, while the other two (ST125-MRSA-IVA and ST146-MRSA-IVA) clones could be derived from the pediatric one. The most striking temporal shift in the dominance of MRSA clones was the replacement of the multidrug-resistant and highly epidemic Iberian clone by the so-called British EMRSA-16 clone during the 5-year surveillance period. Our results are in accordance with previously stated findings showing the worldwide hospital dominance of relatively few pandemic and presumably virulent MRSA clones. We report for the first time the detection in Spain of the British EMRSA-15 and pediatric clones, as well as the abrupt replacement of the Iberian by the EMRSA-16 as the major MRSA clone.Staphylococcus aureus is the causal agent of most staphylococcal pathologies and is currently a versatile microbial pathogen that has evolved resistance to all antibiotic classes. It is associated with serious community-acquired and nosocomial diseases, although most life-threatening S. aureus infections are hospital acquired (4, 8). Its high level of adaptation to hospital environments has been deeply facilitated by the acquisition of methicillin resistance, an evolutionary step that converted S. aureus to methicillin-resistant S. aureus (MRSA), one of the most common nosocomial pathogens nowadays (19). MRSA emerged with the introduction of an exogenous DNA element into its genome, the staphylococcal cassette chromosome mec (SCCmec), which carries the methicillin resistance mecA gene (16). Recent data shows that acquisition of SCCmec has occurred on multiple occasions and that at least five different methicillin-sensitive S. aureus phylogenetic lineages acquired the element (29). Four main structural types of SCCmec, which differ in size and composition, have been described for S. aureus (14, 15, 20).Genetic studies using molecular typing methods have shown that most hospital-acquired MRSA infections worldwide are due to any of the so-called epidemic MRSA (EMRSA). These EMRSA clones present great fitness to hospital environments and, consequently, are established in many hospitals and have spread internationally (7). This situation highlights the importance of monitoring the distribution and routes of dissemination of such EMRSA clones at both levels, within hospitals and between distant locations (24). With this purpose, several molecular techniques and an international common nomenclature have been applied to track EMRSA (10, 27). Pulsed-field gel electrophoresis (PFGE) is considered the “gold standard” for establishing clonal relationships at the local level, but its detection capacity seems to make it also too discriminative for global comparisons (5, 31). By contrast, multilocus sequence typing (MLST) has been verified as an adequate method for long-term and global epidemiological studies (11, 48). Combination of MLST with SCCmec typing permits the unambiguous assignment of collections of MRSA isolates to known or new MRSA clones (10).The aim of this study was to identify MRSA clones circulating in the Nuestra Señora de Candelaria University Hospital (HUNSC), Tenerife, Spain, and to track shifts in their prevalence during a 5-year period (1998 to 2002). For this purpose, we used a combination of different molecular typing methods, including PFGE, MLST, and SCCmec typing.     

Human Cytomegalovirus: detection of congenital and perinatal infection in Argentina

Background  

Human cytomegalovirus (CMV) is one of the most commonly found agents of congenital infections. Primary maternal infection is associated with risk of symptomatic congenital diseases, and high morbidity is frequently associated with very low birth weight. Neonates with asymptomatic infection develop various sequelae during infancy. This is the first Argentine study performed in neonates with congenital and postnatal HCMV infection. The purpose of this study was to evaluate the performance of the polymerase chain reaction (PCR) technique with different pairs of primers, to detect cytomegalovirus isolated in tissue cultures and directly in urine and dried blood spot (DBS) specimens. Results were compared with IgM detection.     

Impairments and activity limitations in subjects with chronic upper-limb complex regional pain syndrome type I

OBJECTIVE: To determine the degree of impairments and activity limitations and their interrelationship in complex regional pain syndrome type I (CRPS type I). DESIGN: Cross-sectional study interrelating impairments and objectively measured activity limitations. SETTING: Ambulatory and home environment. PARTICIPANTS: Thirty nonacute upper-limb CRPS type I subjects. INTERVENTIONS: Not applicable.Main outcome measures Sensory, motor, and autonomic impairments, as well as activity-limitation outcome measures. The latter were objectively measured with a novel upper-limb activity monitor (based on ambulatory accelerometry). RESULTS: All subjects were impaired to some degree but with a large variability with respect to magnitude. Regarding activity limitations, the involved upper limb was clearly less active (lower intensity and percentage of activity) than the noninvolved side. Impaired active range of motion (adjusted R(2) range, 18%-39%) and grip strength (adjusted R(2) range, 12%-45%) were the most important factors explaining variance in activity limitations. CONCLUSIONS: All subjects were still impaired nearly 3 years after the causative event. The involved upper limb was also clearly less active than the noninvolved side, especially when the subjects were sitting and when the dominant side was involved. The more impairments a subject had, especially motor impairments, the more activity limitations were present.     

Effects of long-term exposure to Cu2+ and Cd2+ on the pentose phosphate pathway dehydrogenase activities in the ovary of adult Bufo arenarum: possible role as biomarker for Cu2+ toxicity

The effects of copper and cadmium on metabolism through the pentose phosphate pathway were evaluated in Bufo arenarum toad ovary. The effects of the two metals on dehydrogenases from this pathway were evaluated by three experiments: (1) in samples obtained from control females with addition of the metals to the reaction mixture (in vitro), (2) in samples obtained from control females and after long-term exposure of females to 4 and 100 microg/L of Cu or Cd in the incubation media (in vitro after exposure to the metals in vivo), and (3) 14CO2 production through the pentose phosphate pathway was evaluated after [U-14C]glucose microinjection on ovulated oocytes (in vivo after microinjection of the metals). Results from (1) evidenced inhibition of both enzyme activities but only above 1.5 mM Cu and Cd added to the reaction mixture. In (2) both glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase activities decreased in samples from the ovaries of females exposed in vivo to Cu, in a concentration-dependent manner (up to 90% in females exposed to 100 microg/L Cu: 2.12 +/- 1.57 NADPH micromol/min microg protein x 10(-5) vs 19.97 +/- 8.54 in control females). Cd treatment of the toads only rendered an inhibitory effect on 6-phosphogluconate dehydrogenase activity after exposure to 4 microg/L of the bivalent cation. (3) In vivo 14CO2 evolution significantly decreased in oocytes coinjected with 6.3 x 10(-3) mM Cu (calculated intracellular final concentration of the metal injected) and radioactive glucose. Cu and Cd concentration in samples from exposed females were always under detection limit by particle-induced X-ray emission. The results presented here are in agreement with a role for both glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase activities determination as biomarkers of effect and exposure for Cu but not for Cd toxicity.     

Effect of filgrastim on serum lactate dehydrogenase and alkaline phosphatase values in early breast cancer patients

To improve chemotherapy dose intensity and to optimize the use of granulocyte-colony stimulating factor, 506 patients with early breast cancer were randomly assigned to high dose epirubicin and cyclophospamide (EC) with or without prophylactic subcutaneously filgrastim, according to 5 different schedules: 480 µg or 300 µg daily or every other day, on day 8 through day 14, and 300 µg daily on days 8 and 12 of each chemotherapy course, Serum levels of lactate dehydrogenase (LDH) and alkaline phosphatase (AP) were significantly higher in patients given EC plus filgrastim than EC alone (P + 0.0001), the rate of G1-3 toxicity being 33.4% and 13.1% vs. 1.6% and 1%, respectively. No clinical evidence of filgrastim-related hepatic damage or significant difference in transaminase and γ-GT elevation was seen between the two groups. LDH and AP closely resembled peripheral blood leukocytes count and increased with increasing leucocytosis, throughout the 5 schedules. Although no patient continued treatment for filgrastim-related side effects, and LDH and AP rises resolved spontaneously within 3 weeks following the chemotherapy course, physicians should be aware of the transient and innocuous change in serum chemistry associated to leucocytosis, since it could be misinterpreted as expression of disease activity.     

Oxidative stress markers in aqueous humor of glaucoma patients

PURPOSE: Oxidative stress and antioxidant status in eye tissues may be associated with glaucomatous damage. The aim of this study was to establish the antioxidant status of aqueous humor of patients with primary open-angle glaucoma. For this purpose the authors measured the total reactive antioxidant potential (TRAP) and the activities of the antioxidant enzymes superoxide dismutase (SOD), catalase, and glutathione peroxidase. DESIGN: Case control study. METHODS: Aqueous humor was obtained at the time of surgery from 24 patients with glaucoma and 24 cataract patients; TRAP was measured by chemiluminescence. Activities of the antioxidant enzymes were measured spectrophotometrically. Superoxide dismutase activity was determined by inhibition of the rate of adrenochrome formation at 480 nm. Catalase activity was evaluated by decrease of H(2)O(2) absorbance at 240 nm. Glutathione peroxidase (GPx) activity was determined following nicotinamide adenine dinucleotide phosphate oxidation at 340 nm. RESULTS: Total reactive antioxidant potential value of the cataract group was 124 +/- 5 micromol/l Trolox. This value was significantly decreased, by 64%, in glaucoma patients. An increase of 57% in SOD activity was observed in glaucoma patients when compared with cataract patients (41.7 +/- 2.7 U SOD/ml). Glutathione activity was threefold higher in glaucoma patients than in the cataract group (6.1 +/- 0.6 U/ml). No significant changes were found in catalase levels. CONCLUSIONS: Oxidative stress may lead to an induction of antioxidant enzymes and contribute to TRAP decrease. Superoxide dismutase, GPx activities, and TRAP may be useful oxidative stress markers in aqueous humor of glaucoma patients.     

Lonidamine: efficacy and safety in clinical trials for the treatment of solid tumors

Lonidamine, a derivate of indazole-3-carboxylic acid, is an antineoplastic drug with a typical mechanism of action. Lonidamine has no function on cellular nucleic acids or protein synthesis, whereas it exerts a powerful inhibitory effect on oxygen consumption, aerobic glycolysis and lactate transport and accumulation of neoplastic cells. Nevertheless, its proven ability to modify the permeability of membranes is consistent with the possible increase of drug uptake, reverse of drug resistance and triggering of apoptotic pathway. Lonidamine has been experimentally shown to potentiate the cytotoxic effects of anthracyclines in human breast cancer cell lines and cisplatin activity in both platinum-sensitive and platinum-resistant human ovarian carcinoma cell lines. Since the specific mechanism of action and side effects are not overlapping with those of standard antineoplastic agents, combination of lonidamine with standard chemotherapy has been widely investigated for the treatment of solid tumors. Additionally, the enhancement of radiotherapy activity by lonidamine has been considered for palliative therapy of lesions from metastatic cancers. The encouraging results of phase II-III trials for the treatment of advanced breast, ovarian and lung cancer must be confirmed by larger studies. Specifically designed studies to address the role of lonidamine in the adjuvant setting are warranted. Lonidamine, a dechlorinate derivative of indazole-3-carboxylic acid, has proved to exert a powerful antiproliferative effect and to impair the energy metabolism of neoplastic cells. Herein we review the current experience on combining lonidamine and chemotherapy and/or radiation therapy in the treatment of solid tumors. Several studies have been published on this topic. The total number of trials reported in literature and length of follow-up are still insufficient to draw a firm conclusion. However, the available data demonstrate a significant role of lonidamine in modulating anthracycline and platinum compound activity.