Mutations in aarE, the ubiA Homolog of Providencia stuartii, Result in High-Level Aminoglycoside Resistance and Reduced Expression of the Chromosomal Aminoglycoside 2′-N-Acetyltransferase

The aarE1 allele was identified on the basis of the resulting phenotype of increased aminoglycoside resistance. The aarE1 mutation also resulted in a small-colony phenotype and decreased levels of aac(2′)-Ia mRNA. The deduced AarE gene product displayed 61% amino acid identity to the Escherichia coli UbiA protein, an octaprenyltransferase required for the second step of ubiquinone biosynthesis. Complementation experiments in both Providencia stuartii and E. coli demonstrated that aarE and ubiA are functionally equivalent.     

Mutations Decreasing Intrinsic β-Lactam Resistance Are Linked to Cell Division in the Nosocomial Pathogen Acinetobacter baumannii

Transposon mutagenesis was used to identify novel determinants of intrinsic β-lactam resistance in Acinetobacter baumannii. An EZ-Tn5 transposon insertion in a gene corresponding to the A1S_0225 sequence resulted in a 4-fold decrease in resistance to ampicillin, cefotaxime, imipenem, and ceftriaxone but did not alter resistance to other classes of antibiotics. Based on this phenotype, the gene was designated blhA (β-lactam hypersusceptibility). The blhA::EZ-Tn5 mutation conferred a similar phenotype in A. baumannii strain ATCC 17978. The wild-type blhA gene complemented the blhA::EZTn5 insertion and restored β-lactam resistance levels back to wild-type levels. The blhA mutation also increased β-lactam susceptibility in an adeB adeJ double mutant, indicating that the blhA mutation acted independently of these efflux systems to mediate susceptibility. In addition, mRNA levels for the bla_OXA and bla_ADC β-lactamase genes were not altered by the blhA mutation. The blhA mutation resulted in a prominent cell division and morphological defect, with cells exhibiting a highly elongated phenotype, combined with large bulges in some cells. The blhA gene is unique to Acinetobacter and likely represents a novel gene involved in cell division. Three additional mutations, in zipA, zapA, and ftsK, each of which encode predicted cell division proteins, also conferred increased β-lactam susceptibility, indicating a common link between cell division and intrinsic β-lactam resistance in A. baumannii.     

An extracellular factor regulating expression of the chromosomal aminoglycoside 2'-N-acetyltransferase of Providencia stuartii.

The chromosomal aac(2')-Ia gene in Providencia stuartii encodes a housekeeping 2'-N-acetyltransferase [AAC(2')-Ia] involved in the acetylation of peptidoglycan. In addition, the AAC(2')-Ia enzyme also acetylates and confers resistance to the clinically important aminoglycoside antibiotics gentamicin, tobramycin, and netilmicin. Expression of the aac(2')-Ia gene was found to be strongly influenced by cell density, with a sharp decrease in aac(2')-Ia mRNA accumulation as cells approached stationary phase. This decrease was mediated by the accumulation of an extracellular factor, designated AR (for acetyltransferase repressing)-factor. AR-factor was produced in both minimal and rich media and acted in a manner that was strongly dose dependent. The activity of AR-factor was also pH dependent, with optimal activity at pH 8.0 and above. Biochemical characterization of conditioned media from P. stuartii has shown that AR-factor is between 500 and 1,000 Da in molecular size and is heat stable. In addition, AR-factor was inactivated by a variety of proteases, suggesting that it may be a small peptide.     

Role of ultrasound in detection of radiolucent foreign bodies in extremities

Removal of foreign bodies from soft tissues in emergency is very challenging and becomes more problematic when it is radiolucent. Blind exploration is sometimes hazardous for patients especially when it is in proximity to a vessel or a nerve or an overlying tendon. The purpose of this study was to determine the accuracy of ultrasonography (USG) in detecting radiolucent soft tissue foreign bodies in the extremities. From January 2014 to January 2016, 120 patients with either a positive history or clinically suspected soft tissue foreign body and negative radiography were evaluated by USG with a high-frequency (13–6 MHz) linear-array transducer. The sonographic findings were used to guide surgical exploration. Out of 120 patients who underwent surgical exploration, USG was positive in 114 cases, and foreign body was retrieved in 108 cases, and among the six cases where USG was negative, foreign body was retrieved from one case. In one case with strong clinical suspicion of foreign body USG was falsely negative. Majority of foreign bodies were removed from foot (69 cases) and hands (26 cases), and rest of foreign bodies were removed from ankle (4 cases), wrist (3 cases), thigh (2 cases), leg (1 case), knee (2 cases), forearm (2 cases). Accuracy, sensitivity, and positive predictive value were determined as 94.16, 99.08, and 94.13%, respectively. The real-time high-frequency USG is a highly sensitive and accurate tool for detecting and removing radiolucent foreign bodies which cannot be visualized by routine radiography.     

Glycyrrhizic acid prevents ultraviolet‐B‐induced photodamage: a role for mitogen‐activated protein kinases,nuclear factor kappa B and mitochondrial apoptotic pathway

Glycyrrhizic acid (GA), a natural triterpene, has received attention as an agent that has protective effects against chronic diseases including ultraviolet UV‐B‐induced skin photodamage. However, the mechanism of its protective effect remains elusive. Here, we used an immortalized human keratinocyte cell line (HaCaT) and a small animal model (BALB/c mice), to investigate the protective effects of GA against UV‐B‐induced oxidative damage, and additionally, delineated the molecular mechanisms involved in the UV‐B‐mediated inflammatory and apoptotic response. In the HaCaT cells, GA inhibited the UV‐B‐mediated increase in intracellular reactive oxygen species (ROS) and down‐regulated the release of pro‐inflammatory cytokines interleukin (IL)‐1α, ‐1β and ‐6, tumor necrosis factor (TNF)‐α and prostaglandin E2 (PGE2). GA inhibited UV‐B‐mediated activation of p38 and JNK MAP kinases, COX‐2 expression and nuclear translocation of NF‐κB. Furthermore, GA inhibited UV‐B‐mediated apoptosis by attenuating translocation of Bax from the cytosol to mitochondria, thus preserving mitochondrial integrity. GA‐treated HaCaT cells also exhibited elevated antiapoptotic Bcl‐2 protein, concomitant with reduced caspase‐3 cleavage and decreased PARP‐1 protein. In BALB/c mice, topical application of GA on dorsal skin exposed to UV‐B irradiation protected against epidermal hyperplasia, lymphocyte infiltration and expression of several inflammatory proteins, p38, JNK, COX‐2, NF‐κB and ICAM‐1. Based on the above findings, we conclude that GA protects against UV‐B‐mediated photodamage by inhibiting the signalling cascades triggered by oxidative stress, including MAPK/NF‐κB activation, as well as apoptosis. Thus, GA has strong potential to be used as a therapeutic/cosmeceutical agent against photodamage.     

Analysis of antibiotic resistance genes in multidrug-resistant Acinetobacter sp. isolates from military and civilian patients treated at the Walter Reed Army Medical Center

Military medical facilities treating patients injured in Iraq and Afghanistan have identified a large number of multidrug-resistant (MDR) Acinetobacter baumannii isolates. In order to anticipate the impact of these pathogens on patient care, we analyzed the antibiotic resistance genes responsible for the MDR phenotype in Acinetobacter sp. isolates collected from patients at the Walter Reed Army Medical Center (WRAMC). Susceptibility testing, PCR amplification of the genetic determinants of resistance, and clonality were determined. Seventy-five unique patient isolates were included in this study: 53% were from bloodstream infections, 89% were resistant to at least three classes of antibiotics, and 15% were resistant to all nine antibiotics tested. Thirty-seven percent of the isolates were recovered from patients nosocomially infected or colonized at the WRAMC. Sixteen unique resistance genes or gene families and four mobile genetic elements were detected. In addition, this is the first report of bla(OXA-58)-like and bla(PER)-like genes in the U.S. MDR A. baumannii isolates with at least eight identified resistance determinants were recovered from 49 of the 75 patients. Molecular typing revealed multiple clones, with eight major clonal types being nosocomially acquired and with more than 60% of the isolates being related to three pan-European types. This report gives a "snapshot" of the complex genetic background responsible for antimicrobial resistance in Acinetobacter spp. from the WRAMC. Identifying genes associated with the MDR phenotype and defining patterns of transmission serve as a starting point for devising strategies to limit the clinical impact of these serious infections.     

Helicobacter pylori Infection in Various ABO Blood Groups of Kashmiri Population

Aim: This study was carried out to assess the prevalence of Helicobacter pylori infection in various ABO blood groups of people of Kashmir.Method: The study comprised 80 individuals - 50 peptic ulcer patients (whose disease was diagnosed by endoscopy) and 30 asymptomatic volunteers. Every subject's blood group and Rhesus status was determined by standard serological tests. Helicobacter pylori infection was diagnosed by three different methods viz., one minute endoscopy room test (urease test), Gram staining and by histology. The detection of Helicobacter pylori by histological examination using Giemsa staining was taken as the 'gold standard' for the presence of Helicobacter pylori infection.Results: Out of 80 individuals, 67 were males and 13 females aged between 18-65 years. The majority of peptic ulcer patients had blood group 'O' (n = 28.56%). The prevalence of Helicobacter pylori infection amongst peptic ulcer patients was 76%. There was no difference in Helicobacter pylori positivity in various blood groups.Conclusion: Blood group 'O' though a risk factor for peptic ulcer (Duodenal ulcer) is not a risk factor for acquiring Helicobacter pylori infection.