OBJECTIVETo evaluate the free radical generation and antioxidant enzymes status in murine peritoneal macrophage during in vitro amikacin resistant Pseudomonas aeruginosa (ARPA) treatment with different time interval.

METHODSPeritoneal macrophages were treated with 1×10(8) CFU/mL ARPA cell suspension in vitro for different time interval (1, 2, 3, 6, 12, and 24 h) and super oxide anion generation, NO generation, reduced glutathione level and antioxidant enzymes status were analyzed.

RESULTSSuper oxide anion generation and NO generation got peak at 12 h, indicating maximal free radical generation through activation of NADPH oxidase in murine peritoneal macrophages during ARPA transfection. Reduced glutathione level and antioxidant enzymes status were decreased significantly (P<0.05) with increasing time of ARPA transfection. All the changes in peritoneal macrophages after 12 h in vitro ARPA transfection had significant difference (P<0.05).

CONCLUSIONSFrom this study, it may be summarized that in vitro ARPA infection not only generates excess free radical but also affects the antioxidant system and glutathione cycle in murine peritoneal macrophage.

Amikacin ; pharmacology ; Animals ; Anti-Bacterial Agents ; pharmacology ; Antioxidants ; analysis ; Cells, Cultured ; Drug Resistance, Bacterial ; Free Radicals ; analysis ; Glutathione ; analysis ; Macrophages, Peritoneal ; immunology ; microbiology ; physiology ; Male ; Mice ; Oxidative Stress ; Pseudomonas aeruginosa ; growth & development ; immunology ; Time Factors

AIMTo elucidate the genetic basis for the pronounced resistance that the oral pathogen, Porphyromonas gingivalis (P. gingivalis), exhibits towards the cationic antimicrobial peptide, polymyxin B.

METHODOLOGYA genetic screen of P. gingivalis clones generated by a Tn4400'-based random insertion mutagenesis strategy was performed to identify bacteria harboring novel genetic mutations that render P. gingivalis susceptible to killing by the cationic antimicrobial peptide, polymyxin B (PMB, 50 microg x mL(-1)).

RESULTSP. gingivalis (ATCC 33277) is unusually resistant to the cationic antimicrobial peptide, PMB at relatively high concentrations (200 microg x mL(-1)). Approximately 2,700 independent Tn4400'-derived mutants of P. gingivalis were examined for increased sensitivity to PMB killing at a relatively low dose (50 microg x mL(-1)). A single PMB-sensitive mutant was obtained in this phenotypic screen. We determined that the Tn4400' transposon was integrated into the gene encoding the lipid A 4'-phosphatase, PGN_0524, demonstrating that this insertion event was responsible for its increased susceptibility of this clone to PMB-dependent killing. The resulting mutant strain, designated 0524-Tn4400', was highly sensitive to PMB killing relative to wild-type P. gingivalis, and exhibited the same sensitivity as the previously characterized strain, 0524KO, which bears a genetically engineered deletion in the PGN_0524 locus. Positive ion mass spectrometric structural (MALDI-TOF MS) analyses revealed that lipid A isolates from 0524-Tn4400' and 0524KO strains displayed strikingly similar MALDI-TOF MS spectra that were substantially different from the wildtype P. gingivalis lipid A spectrum. Finally, intact 0524-Tn4400' and 0524KO mutant bacteria, as well as their corresponding LPS isolates, were significantly more potent in stimulating Toll-like receptor 4 (TLR4)-dependent E-selectin expression in human endothelial cells relative to intact wild-type P. gingivalis or its corresponding LPS isolate.

CONCLUSIONThe combined molecular evidence provided in this report suggests that PGN_0524, a lipid A 4'-phosphatase, is the sole genetic element conferring the ability of the periodontopathogen, P. gingivalis, to evade the killing activity of cationic antimicrobial peptides, such as PMB. These data strongly implicate PGN_0524 as a critical virulence factor for the ability of P. gingivalis to evade front-line host innate defenses that are dependent upon cationic antimicrobial peptide activity and TLR 4 sensing.

Anti-Bacterial Agents ; pharmacology ; Chromosome Mapping ; DNA Transposable Elements ; genetics ; Drug Resistance, Bacterial ; genetics ; E-Selectin ; analysis ; immunology ; Endothelial Cells ; immunology ; microbiology ; Gene Deletion ; Humans ; Lipid A ; analysis ; immunology ; Lipopolysaccharides ; analysis ; immunology ; Mutagenesis, Insertional ; genetics ; Open Reading Frames ; genetics ; Phosphoric Monoester Hydrolases ; genetics ; physiology ; Polymyxin B ; pharmacology ; Porphyromonas gingivalis ; enzymology ; genetics ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Toll-Like Receptor 4 ; analysis ; immunology ; Virulence Factors ; physiology

BACKGROUND: Streptococcus pneumoniae causes life-threatening infections such as meningitis, pneumonia, and febrile bacteremia, particularly in young children. The increasing number of drug-resistant isolates has highlighted the necessity for intervening and controlling disease. To achieve this, information is needed on serotype distribution and patterns of antibiotic resistance in children. METHODS: All cases of invasive pneumococcal disease (IPD) in children aged less than 15 yr recorded at King Khalid University Hospital, King Saud University, Riyadh, Saudi Arabia, were reviewed for serotyping and antibiotic susceptibility. Isolates were collected from 78 consecutive patients with IPD between 2009 and 2012. All collected isolates were subjected to serotyping by co-agglutination, sequential multiplex PCR, and single PCR sequetyping as previously described. RESULTS: The most frequently isolated IPD serotypes were 23F, 6B, 19F, 18C, 4, 14, and 19A, which are listed in decreasing order and cover 77% of total isolates. The serotype coverage for the pneumococcal conjugate vaccine (PCV)7, PCV10, and PCV13 was 77%, 81%, and 90%, respectively. Results from sequential multiplex PCR agreed with co-agglutination results. All serotypes could not be correctly identified using single PCR sequetyping. Minimum inhibitory concentration showed that 50 (64%) isolates were susceptible to penicillin, whereas 70 (90%) isolates were susceptible to cefotaxime. CONCLUSIONS: The most common pneumococcal serotypes occur with frequencies similar to those found in countries where the PCV has been introduced. The most common serotypes in this study are included in the PCVs. Addition of 23A and 15 to the vaccine would improve the PCV performance in IPD prevention.
Adolescent ; Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/genetics ; Cefotaxime/pharmacology ; Child ; Child, Preschool ; DNA, Bacterial/analysis ; Humans ; Infant ; Meningitis/*diagnosis/microbiology ; Microbial Sensitivity Tests ; Multiplex Polymerase Chain Reaction ; Penicillins/pharmacology ; Pneumococcal Vaccines/immunology ; Pneumonia/*diagnosis/microbiology ; Protein Tyrosine Phosphatases/genetics ; Retrospective Studies ; Saudi Arabia ; Serotyping ; Streptococcus pneumoniae/*drug effects/genetics

Exposure to cephalosporins could cause occupational allergic diseases in health care workers (HCWs). We evaluated the prevalence of serum specific IgE and IgG antibodies to cephalosporin-human serum albumin (HSA) conjugate and to identify potential genetic risk factors associated with sensitization to cephalosporins in exposed HCWs. The study population consisted of 153 HCWs who had been exposed to antibiotics in a single university hospital and 86 unexposed healthy controls. A questionnaire survey of work-related symptoms (WRS) was administered. A skin-prick test (SPT) was performed, and serum-specific IgE and IgG antibodies to 3 commonly prescribed cephalosporins were measured by ELISA. Four single-nucleotide polymorphisms of the candidate genes related to IgE sensitization were genotyped. The prevalence of WRS to cephalosporins was 2.6%. The prevalence rates of serum-specific IgE and IgG antibodies to cephalosporins were 20.3% and 14.7%, respectively. The FcepsilonR1beta-109T > C polymorphism was significantly associated with IgE sensitization to cephalosporins in HCWs (P = 0.036, OR = 3.553; CI, 1.324-9.532). The in vitro functional assay demonstrated that the T allele of FcepsilonR1beta-109T had greater promoter activity than did the C allele (P < 0.001). The FcepsilonR1beta-109T > C polymorphism may be a potential genetic risk factor for increased IgE sensitization to cephalosporins.
Adult ; Alleles ; Anti-Bacterial Agents/analysis/*immunology ; Cephalosporins/analysis/*immunology ; Enzyme-Linked Immunosorbent Assay ; Female ; Genetic Predisposition to Disease ; Health Personnel ; Humans ; Hypersensitivity/*diagnosis/epidemiology ; Immunoglobulin E/blood ; Immunoglobulin G/blood ; Male ; Occupational Diseases/*chemically induced/epidemiology ; Occupational Exposure ; Odds Ratio ; Questionnaires ; Receptors, IgE/genetics ; Skin Tests ; Young Adult

There are limited data evaluating the relationship between influenza treatment and hospitalization duration. Our purpose assessed the association between different treatments and hospital stay among Korean pediatric influenza patients. Total 770 children < or = 15 yr-of-age hospitalized with community-acquired laboratory-confirmed influenza at three large urban tertiary care hospitals were identified through a retrospective medical chart review. Demographic, clinical, and cost data were extracted and a multivariable linear regression model was used to assess the associations between influenza treatment types and hospital stay. Overall, there were 81% of the patients hospitalized with laboratory-confirmed influenza who received antibiotic monotherapy whereas only 4% of the patients received oseltamivir monotherapy. The mean treatment-related charges for hospitalizations treated with antibiotics, alone or with oseltamivir, were significantly higher than those treated with oseltamivir-only (P < 0.001). Influenza patients treated with antibiotics-only and antibiotics/oseltamivir combination therapy showed 44.9% and 28.2%, respectively, longer duration of hospitalization compared to those treated with oseltamivir-only. Patients treated with antibiotics, alone or combined with oseltamivir, were associated with longer hospitalization and significantly higher medical charges, compared to patients treated with oseltamivir alone. In Korea, there is a need for more judicious use of antibiotics, appropriate use of influenza rapid testing.
Adolescent ; Anti-Bacterial Agents/*therapeutic use ; Antigens, Viral/analysis/immunology ; Antiviral Agents/*therapeutic use ; Child ; Child, Preschool ; Cohort Studies ; Demography ; Drug Therapy, Combination ; Female ; Hospitalization ; Humans ; Infant ; Infant, Newborn ; Influenza A virus/metabolism ; Influenza B virus/metabolism ; Influenza, Human/*drug therapy ; Male ; Oseltamivir/*therapeutic use ; Republic of Korea ; Retrospective Studies

Amoebic liver abscess is a common disease, especially in endemic areas, but it is a rare cause of inferior vena cava (IVC) obstruction, with only a few cases appearing in the literature. We report three cases of amoebic liver abscess complicated with obstruction of the IVC and which responded to conservative treatment or radiological intervention.
Anti-Bacterial Agents/therapeutic use ; Antibodies, Protozoan/analysis ; Entamoeba/immunology/isolation & purification ; Enzyme-Linked Immunosorbent Assay ; Humans ; Liver Abscess, Amebic/complications/*diagnosis/ultrasonography ; Magnetic Resonance Imaging ; Male ; Middle Aged ; Thrombosis/diagnosis/etiology ; Tomography, X-Ray Computed ; Vascular Diseases/*etiology ; *Vena Cava, Inferior ; Young Adult