RAW TROPICAL OYSTERS AS VEHICLES FOR MULTIDRUG-RESISTANT Vibrio parahaemolyticus

The following study aimed to determine the antimicrobial susceptibility profile of Vibrio parahaemolyticus strains from fresh and frozen oysters Crassostrea rhizophorae sold in Fortaleza-Brazil. An antibiogram was performed on 87 isolates using nine antibiotics: gentamicin (Gen 10 µg), ampicillin (Amp 10 µg), penicillin G (Pen 10U), ciprofloxacin (Cip 5 µg), chloramphenicol (Chl 30 µg), nalidixic acid (Nal 30 µg), tetracycline (Tet 30 µg), vancomycin (Van 30 µg) and erythromycin (Ery 15 µg). All strains were resistant to at least one antibiotic, and 85 (97.7%) were multi-resistant, with predominance of the Van+ Pen+Amp resistance profile (n = 46). Plasmid resistance to Pen, Amp and Ery was detected. Thus, the risk that raw oyster consumption poses to the health of consumers is highlighted, due to the fact that these bivalves may host antibacterial-resistant microorganisms.

Gold nanoprobes for the detection of mutations associated with antibiotic resistance in Mycobacterium tuberculosis complex

Dissertação para obtenção do Grau de Mestre em Genética Molecular e Biomedicina

High prevalence of methicillin-resistant Staphylococcus aureus with SCCmec type III in cystic fibrosis patients in southern, Brazil

INTRODUCTION: Bacterial colonization of the lungs is the main cause of morbidity in cystic fibrosis (CF). Pathogens such as Staphylococcus aureus are very well adapted to the pulmonary environment and may persist for years in the same patient. Genetic determinants of these bacteria, such as the presence of SCCmec have recently emerged as a problem in this population of patients. METHODS: Staphylococcus aureus isolates obtained from different clinical materials coming from CF and non-CF patients attended at a cystic fibrosis reference hospital were compared according to SCCmec type and antibiotic susceptibility profile. RESULTS: Three hundred and sixty-four single-patient Staphylococcus aureus isolates were collected, of which 164 (45%) were from CF patients. Among the latter, 57/164 (44.5%) were MRSA, and among the non-CF patients, 89/200 (35%) were MRSA. Associated pathogens were found in 38 CF patients. All 57 MRSA from CF patients harbored the multiresistant cassette type III. In contrast, 31/89 MRSA from non-CF patients harbored SCCmec type I (35%) and 44/89 harbored type III (49%). The antibiotic susceptibility pattern was similar between CF and non-CF patients. CONCLUSIONS: The high prevalence of multiresistant SCCmec type III among CF patients compared with non-CF patients in our institution may make it difficult to control disease progression through antibiotic therapy for promoting the survival of this kind of patient.

Relationship between antibiotic consumption, oropharyngeal colonization, and ventilator-associated pneumonia by Staphylococcus aureus in an intensive care unit of a Brazilian teaching hospital

INTRODUCTION: his study evaluated the consumption of major classes of antibiotics, the colonization of the oropharynx of patients on mechanical ventilation, and the risk of ventilator-associated pneumonia (VAP) caused by Staphylococcus aureus in an intensive care unit for adults. METHODS: A case-control study was carried out using colonized patients (cases) by oxacillin-resistant S. aureus (ORSA) and (controls) oxacillin-sensitive S. aureus (OSSA) from May 2009 to August 2010. The occurrence of VAP by S. aureus was also evaluated in the same period. Antibiotic consumption was expressed as the number of defined daily doses (DDD)/1,000 patient-days for glycopeptides, carbapenems, and extended-spectrum cephalosporins. RESULTS: Three hundred forty-six (56.1%) patients underwent mechanical ventilation with a frequency of oropharyngeal colonization of 36.4%, corresponding to 63.5% for ORSA and 36.5% for OSSA. The risk of illness for this organism was significant (p<0.05), regardless of whether colonization/infection was by ORSA or OSSA. The consumption of antibiotics was high, mainly for broad-spectrum cephalosporins (551.26 DDDs/1,000 patient-days). The high density of use of glycopeptides (269.56 DDDs/1,000 patient-days) was related to colonization by ORSA (Pearson r=0.57/p=0.02). Additionally, age >60 years, previous antibiotic therapy, and previous use of carbapenems were statistically significant by multivariate analysis. CONCLUSIONS: There was a significant relationship between the colonization of the oropharyngeal mucosa and the risk of VAP by both phenotypes. The use of glycopeptides was related to colonization by ORSA.

First case of Helicobacter pyloriinfection resistant to seven antibiotics in Iran

Treatment of Helicobacter pylori infection with common antibiotics is typically recommended for several digestive conditions, including peptic ulcers. However, reports of resistant H. pylori isolates are increasing, and unfortunately, these do not respond to currently available therapeutic regimens. We report the case of a 31-year-old woman with two peptic ulcers in the duodenal antrum. An H. pylori strain was isolated, and tested for antibiotic resistance using agar dilution and disk diffusion. The isolated strain was found to be resistant to all seven antibiotics that were tested. Therefore, constant monitoring for antibiotic resistance should be performed prior to initiating antibiotic therapy.

Aerobic bacterial profile and antibiotic resistance in patients with diabetic foot infections

ABSTRACTINTRODUCTION: This study aimed to determine the frequencies of bacterial isolates cultured from diabetic foot infections and assess their resistance and susceptibility to commonly used antibiotics.METHODS: This prospective study included 41 patients with diabetic foot lesions. Bacteria were isolated from foot lesions, and their antibiotic susceptibility pattern was determined using the Kirby-Bauer disk diffusion method and/or broth method [minimum inhibitory concentration (MIC)].RESULTS: The most common location of ulceration was the toe (54%), followed by the plantar surface (27%) and dorsal portion (19%). A total of 89 bacterial isolates were obtained from 30 patients. The infections were predominantly due to Gram-positive bacteria and polymicrobial bacteremia. The most commonly isolated Gram-positive bacteria were Staphylococcus aureus, followed by Staphylococcus saprophyticus, Staphylococcus epidermidis, Streptococcus agalactiae, and Streptococcus pneumoniae. The most commonly isolated Gram-negative bacteria were Proteus spp. and Enterobacterspp., followed by Escherichia coli, Pseudomonasspp., and Citrobacterspp. Nine cases of methicillin-resistant Staphylococcus aureus (MRSA) had cefoxitin resistance, and among these MRSA isolates, 3 were resistant to vancomycin with the MIC technique. The antibiotic imipenem was the most effective against both Gram-positive and Gram-negative bacteria, and gentamicin was effective against Gram-negative bacteria.CONCLUSIONS: The present study confirmed the high prevalence of multidrug-resistant pathogens in diabetic foot ulcers. It is necessary to evaluate the different microorganisms infecting the wound and to know the antibiotic susceptibility patterns of the isolates from the infected wound. This knowledge is crucial for planning treatment with the appropriate antibiotics, reducing resistance patterns, and minimizing healthcare costs.

High-throughput hydrophilic interaction chromatography coupled to tandem mass spectrometry for the optimized quantification of the anti-Gram-negatives antibiotic colistin A/B and its pro-drug colistimethate.

Colistin is a last resort's antibacterial treatment in critically ill patients with multi-drug resistant Gram-negative infections. As appropriate colistin exposure is the key for maximizing efficacy while minimizing toxicity, individualized dosing optimization guided by therapeutic drug monitoring is a top clinical priority. Objective of the present work was to develop a rapid and robust HPLC-MS/MS assay for quantification of colistin plasma concentrations. This novel methodology validated according to international standards simultaneously quantifies the microbiologically active compounds colistin A and B, plus the pro-drug colistin methanesulfonate (colistimethate, CMS). 96-well micro-Elution SPE on Oasis Hydrophilic-Lipophilic-Balanced (HLB) followed by direct analysis by Hydrophilic Interaction Liquid Chromatography (HILIC) with Ethylene Bridged Hybrid - BEH - Amide phase column coupled to tandem mass spectrometry allows a high-throughput with no significant matrix effect. The technique is highly sensitive (limit of quantification 0.014 and 0.006μg/mL for colistin A and B), precise (intra-/inter-assay CV 0.6-8.4%) and accurate (intra-/inter-assay deviation from nominal concentrations -4.4 to +6.3%) over the clinically relevant analytical range 0.05-20μg/mL. Colistin A and B in plasma and whole blood samples are reliably quantified over 48h at room temperature and at +4°C (<6% deviation from nominal values) and after three freeze-thaw cycles. Colistimethate acidic hydrolysis (1M H2SO4) to colistin A and B in plasma was completed in vitro after 15min of sonication while the pro-drug hydrolyzed spontaneously in plasma ex vivo after 4h at room temperature: this information is of utmost importance for interpretation of analytical results. Quantification is precise and accurate when using serum, citrated or EDTA plasma as biological matrix, while use of heparin plasma is not appropriate. This new analytical technique providing optimized quantification in real-life conditions of the microbiologically active compounds colistin A and B offers a highly efficient tool for routine therapeutic drug monitoring aimed at individualizing drug dosing against life-threatening infections.

A quorum sensing small volatile molecule promotes antibiotic tolerance in bacteria.

Bacteria can be refractory to antibiotics due to a sub-population of dormant cells, called persisters that are highly tolerant to antibiotic exposure. The low frequency and transience of the antibiotic tolerant "persister" trait has complicated elucidation of the mechanism that controls antibiotic tolerance. In this study, we show that 2' Amino-acetophenone (2-AA), a poorly studied but diagnostically important small, volatile molecule produced by the recalcitrant gram-negative human pathogen Pseudomonas aeruginosa, promotes antibiotic tolerance in response to quorum-sensing (QS) signaling. Our results show that 2-AA mediated persister cell accumulation occurs via alteration of the expression of genes involved in the translational capacity of the cell, including almost all ribosomal protein genes and other translation-related factors. That 2-AA promotes persisters formation also in other emerging multi-drug resistant pathogens, including the non 2-AA producer Acinetobacter baumannii implies that 2-AA may play an important role in the ability of gram-negative bacteria to tolerate antibiotic treatments in polymicrobial infections. Given that the synthesis, excretion and uptake of QS small molecules is a common hallmark of prokaryotes, together with the fact that the translational machinery is highly conserved, we posit that modulation of the translational capacity of the cell via QS molecules, may be a general, widely distributed mechanism that promotes antibiotic tolerance among prokaryotes.

Y-688, a new quinolone active against quinolone-resistant Staphylococcus aureus: lack of in vivo efficacy in experimental endocarditis.

Y-688 is a new fluoroquinolone with increased activity against ciprofloxacin-resistant staphylococci. The MICs of Y-688 and other quinolones were determined for 58 isolates of ciprofloxacin-resistant and methicillin-resistant Staphylococcus aureus (MRSA). The MICs at which 50% and 90% of bacteria were inhibited were &gt;/=128 and &gt;/=128 mg/liter, respectively, for ciprofloxacin, 16 and 32 mg/liter, respectively, for sparfloxacin, and 0.25 and 1 mg/liter, respectively, for Y-688. This new quinolone was further tested in rats with experimental endocarditis due to either of two isolates of ciprofloxacin-resistant MRSA (namely, P8/128 and CR1). Infected animals were treated for 3 days with ciprofloxacin, vancomycin, or Y-688. Antibiotics were administered through a computerized pump to simulate human-like pharmacokinetics in the serum of rats. The anticipated peak and trough levels of Y-688 were 4 and 1 mg/liter at 0.5 and 12 h, respectively. Treatment with ciprofloxacin was ineffective. Vancomycin significantly decreased vegetation bacterial counts for both organisms (P less, similar 0.05). In contrast, Y-688 only marginally decreased vegetation bacterial counts (P greater, similar 0.05). Moreover, several vegetation that failed Y-688 treatment grew staphylococci for which the MICs of the test antibiotic were increased two to eight times. Y-688 also selected for resistance in vitro, and isolates for which the MICs were increased eight times emerged at a frequency of ca. 10(-8). Thus, in spite of its low MIC for ciprofloxacin-resistant MRSA, Y-688 failed in vivo and its use carried the risk of resistance selection. The fact that ciprofloxacin-resistant staphylococci became rapidly resistant to this potent new drug suggests that the treatment of ciprofloxacin-resistant MRSA with new quinolones might be more problematic than expected.

Parenteral sparfloxacin compared with ceftriaxone in treatment of experimental endocarditis due to penicillin-susceptible and -resistant streptococci.

A new, investigational, parenteral form of sparfloxacin was compared with ceftriaxone in the treatment of experimental endocarditis caused by either of three penicillin-susceptible streptococci or one penicillin-resistant streptococcus. Both drugs have prolonged half-lives in serum, allowing single daily administration to humans. Sparfloxacin had relatively low MICs (0.25 to 0.5 mg/liter) for all four organisms and was also greater than or equal to eight times more effective than the other quinolones against 21 additional streptococcal isolates recovered from patients with bacteremia. Ceftriaxone MICs were 0.032 to 0.064 mg/liter for the penicillin-susceptible strains and 2 mg/liter for the resistant isolate. Both antibiotics resulted in moderate bacterial killing in vitro. Rats with catheter-induced aortic vegetations were inoculated with 10(7) CFU of the test organisms. Antibiotic treatment was started 48 h later and lasted either 3 or 5 days. The drugs were injected at doses which mimicked the kinetics in human serum produced by one intravenous injection of 400 mg of sparfloxacin (i.e., the daily dose expected to be given to human adults) and 2 g of ceftriaxone. Both antibiotics significantly decreased the bacterial densities in the vegetations. However, sparfloxacin was slower than ceftriaxone in its ability to eradicate valvular infection caused by penicillin-susceptible bacteria. While this difference was quite marked after 3 days of therapy, it tended to vanish when treatment was prolonged to 5 days. In contrast, sparfloxacin was very effective against the penicillin-resistant isolate, an organism against which ceftriaxone therapy failed in vivo. No sparfloxacin-resistant mutant was selected during therapy. Thus, in the present experimental setting, this new, investigational, parenteral form of sparfloxacin was effective against severe infections caused by both penicillin-susceptible and penicillin-resistant streptococci.

Management of a post-operative multi-resistant infectious spondylitis associated with a kyphotic deformity.

Anterior spinal infection (prevertebral abscess and/or discitis) after posterior instrumentation for vertebral fractures is a challenging complication, since a new implant may become necessary anteriorly, in a septic environment. Generally accepted management guidelines are yet to be established. The authors present a case of posterior instrumentation for fractures of T12 and L1, complicated after 9 months with an anterior infection (prevertebral abscess and discitis) with extended-spectrum beta-lactamase (ESBL) producing Escherichia coli (E. coli). This case is unique in that the multi-resistant organism was isolated only after the second stage of infection treatment, which consisted of anterior débridement and anterior implantation of titanium cages and rods. In this particular case, infection was controlled despite implantation of multiple cages, screws and rods, and fusion was achieved, by means of intravenous antibiotic treatment for 12 months. At the latest follow-up, 24 months post surgery, there was no evidence of infection. This problem case may be helpful for surgeons confronted with spinal deformities secondary to infections with multi-resistant organisms.

Slime production and antibiotic susceptibility in staphylococci isolated from clinical samples

A total of 187 isolates from several clinical specimens were identified to species level as 129 Staphylococcus aureus strains and 58 coagulase-negative staphylococci (CNS) strains by the API Staph System (Biomerieux). Slime production was detected both by the conventional Christensen's method as well as by the Congo red agar method. Seventy-two strains of staphylococci isolates (38.5%) were found to be slime producers by Christensen's test tube method whereas 58 strains (31%) were slime positive with Congo red agar method. There was no statistically significant difference between the two methods for the detection of slime production (P > 0.05). Susceptibility of isolates against antimicrobial agents was tested by the disk diffusion method. Staphylococcal species had resistance to one or more antibiotics. Among the various antimicrobial agents, oxacillin (71.1%) and erythromycin (47.1%) showed higher resistance than most of the agents used against all isolates. Oxacillin resistant S. aureus (ORSA) and oxacillin resistant coagulase-negative staphylococci (ORCNS), 97 (75.2%) and 36 (62.1%) respectively were frequently observed in strains isolated from clinical materials. Among the ORSA strains, two strains were resistant to vancomycin. Moreover, 96 (74.4%) of 129 S. aureus strains were positive for blactamase enzyme. However, 78 (81.25%) of 96 b-lactamase positive S. aureus strains were b-lactamase positive ORSA isolates, but none of them had vancomycin resistance.

Antibiotic susceptibility of Waddlia chondrophila in Acanthamoeba castellanii amoebae.

Waddlia chondrophila is an emerging cause of miscarriage in bovines and humans. Given the strict intracellular growth of this Chlamydia-like organism, its antibiotic susceptibility was tested by amoebal coculture, cell culture, and real-time PCR. W. chondrophila was susceptible to doxycycline and azithromycin but resistant to beta-lactams and fluoroquinolones.

Nosocomial Outbreak of Infection With Pan-Drug-Resistant Acinetobacter baumannii in a Tertiary Care University Hospital

OBJECTIVE To describe what is, to our knowledge, the first nosocomial outbreak of infection with pan-drug-resistant (including colistin-resistant) Acinetobacter baumannii, to determine the risk factors associated with these types of infections, and to determine their clinical impact. DESIGN Nested case-control cohort study and a clinical-microbiological study. SETTING A 1,521-bed tertiary care university hospital in Seville, Spain. PATIENTS Case patients were inpatients who had a pan-drug-resistant A. baumannii isolate recovered from a clinical or surveillance sample obtained at least 48 hours after admission to an intensive care unit (ICU) during the time of the epidemic outbreak. Control patients were patients who were admitted to any of the "boxes" (ie, rooms that partition off a distinct area for a patient's bed and the equipment needed to care for the patient) of an ICU for at least 48 hours during the time of the epidemic outbreak. RESULTS All the clinical isolates had similar antibiotic susceptibility patterns (ie, they were resistant to all the antibiotics tested, including colistin), and, on the basis of repetitive extragenic palindromic-polymerase chain reaction, it was determined that all of them were of the same clone. The previous use of quinolones and glycopeptides and an ICU stay were associated with the acquisition of infection or colonization with pan-drug-resistant A. baumannii. To control this outbreak, we implemented the following multicomponent intervention program: the performance of environmental decontamination of the ICUs involved, an environmental survey, a revision of cleaning protocols, active surveillance for colonization with pan-drug-resistant A. baumannii, educational programs for the staff, and the display of posters that illustrate contact isolation measures and antimicrobial use recommendations. CONCLUSIONS We were not able to identify the common source for these cases of infection, but the adopted measures have proven to be effective at controlling the outbreak.

Streptococcus pneumoniae bacteremia: clinical and microbiological epidemiology in a health area of Southern Spain

Streptococcus pneumoniae remains an important cause of bacteremia worldwide. Last years, a decrease of S. pneumoniae penicillin-resistant isolates has been observed. The objective of this study was to describe the episodes of bacteremia due to S. pneumoniae during a period of 11 years. Epidemiological and clinical data, serotypes causing bacteremia, antibiotic susceptibility and prognosis factors were studied. Over a period of 11 years, all the episodes of S. pneumoniae bacteremia were analysed. Their clinical and microbiological features were recorded. Statistical analysis was carried out to determine risk factors for pneumococcal bacteremia and predictors of fatal outcome. Finally, 67 S. pneumoniae bacteremia episodes were included in this study. The majority of cases were produced in white men in the middle age of their life. The main predisposing factors observed were smoking, antimicrobial and/or corticosteroids administration, chronic pulmonary obstructive disease and HIV infection, and the most common source of bacteremia was the low respiratory tract. The main serotypes found were 19A, 1, 14 and 7F. Seventy-seven percent of these isolates were penicillin-susceptible, and the mortality in this serie was really low. Statistical significance was observed between age, sex and race factors and the presence of bacteremia, and there was relationship between the patient’s condition and the outcome. In our study, S. pneumoniae bacteremia is mainly from community-acquired origin mainly caused in men in the median age of the life. 40% of bacteremias were caused by serotypes 19A, 1, 7F and 14. During the period of study the incidence of bacteremia was stable and the mortality rate was very low.