Prevalence and determinants of methicillin-resistant Staphylococcus aureus and methicillin-sensitive Staphylococcus aureus carrier among staff of a tertiary eye hospital in Saudi Arabia.

OBJECTIVES: The objective of this study was to estimate the prevalence and risk factors of methicillin-sensitive Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) carriers among staff of a tertiary eye hospital in Saudi Arabia. METHODS: This retrospective study was conducted in 2019. Nasal and axillary swabs of health-care staff were used to determine carriers of MRSA. Bacteria were identified by culture and sensitivity tests. These isolates were grouped as antibiotic resistant, sensitive, and others not in the S. aureus group. Demographics and other determinants were associated with carrier status. RESULTS: We evaluated the carrier status of 430 staff. There were 110 (24.9%; 95% confidence interval [CI]: 21.5, 29.7) S. aureus-positive staff, 21 (11.7%; 95% CI: 11.7, 26.4) of who carried the MRSA strain. Carrier status was significantly higher among physicians (31%) compared to nurses (22.5%) and other staff (5.7%) (P < 0.001). MRSA carrier status was significantly associated with >5 years of employment at the eye hospital (P = 0.02). MRSA was significantly associated with staff who were of Indian nationality (75%) compared to other nationalities (P = 0.04) and those who were at the hospital for <5-year stay compared more than 5 years at the hospital (P = 0.001). All carriers responded to decolonization treatment. CONCLUSIONS: The high prevalence of MRSA and relatively easy treat MRSA carriers points at the need for universal screening for MRSA carriers among eye health staff.

Response to Comments on "Designing river flows to improve food security futures in the Lower Mekong Basin".

Sabo et al presented an empirically derived algorithm defining the socioecological response of the Tonle Sap Dai fishery in the Cambodian Mekong to basin-scale variation in hydrologic flow regime. Williams suggests that the analysis leading to the algorithm is flawed because of the large distance between the gauge used to measure water levels (hydrology) and the site of harvest for the fishery. Halls and Moyle argue that Sabo et al's findings are well known and contend that the algorithm is not a comprehensive assessment of sustainability. We argue that Williams' critique stems from a misunderstanding about our analysis; further clarification of the analysis is provided. We regret not citing more of the work indicated by Halls and Moyle, yet we note that our empirical analysis provides additional new insights into Mekong flow-fishery relationships.

Manual red cell exchange transfusion to avert sickle cell related complications.

Sickle cell-beta thalassemia is a double heterozygous state. Red cell exchange (RCE) transfusion reduces the concentration of sickle cells without increasing the hematocrit or whole-blood viscosity. It can be performed manually or by erythrocytapheresis. RCE transfusion is an effective tool for both acute and chronic complications of sickle cell disease. In patients unaffording erythrocytapheresis, even manual RCE can give favorable results. A 37-year-old male, a known case of sickle cell-beta+ thalassemia (ßsß+), presented with avascular necrosis of right femur and humeral head. He was posted for the right hip arthroplasty and shoulder hemiarthroplasty. Successful manual RCE transfusions were done. The hemoglobin S levels decreased postmanual RCE procedures, and the patient was operated successfully.

Designing river flows to improve food security futures in the Lower Mekong Basin.

Rivers provide unrivaled opportunity for clean energy via hydropower, but little is known about the potential impact of dam-building on the food security these rivers provide. In tropical rivers, rainfall drives a periodic flood pulse fueling fish production and delivering nutrition to more than 150 million people worldwide. Hydropower will modulate this flood pulse, thereby threatening food security. We identified variance components of the Mekong River flood pulse that predict yield in one of the largest freshwater fisheries in the world. We used these variance components to design an algorithm for a managed hydrograph to explore future yields. This algorithm mimics attributes of discharge variance that drive fishery yield: prolonged low flows followed by a short flood pulse. Designed flows increased yield by a factor of 3.7 relative to historical hydrology. Managing desired components of discharge variance will lead to greater efficiency in the Lower Mekong Basin food system.

Vision Screening of Ophthalmic Nursing Staff in a Tertiary Eye Care Hospital: Outcomes and ocular healthcare-seeking behaviours.

OBJECTIVES: This study aimed to evaluate ocular healthcare-seeking behaviours and vision screening outcomes of nursing staff at a tertiary eye care hospital. METHODS: This study was conducted between April and September 2016 among all 500 nurses employed at the King Khaled Eye Specialist Hospital, Riyadh, Saudi Arabia. Data were collected on age, gender, use of visual aids, the presence of diabetes, a history of refractive surgery and date of last ocular health check-up. Participants were tested using a handheld Spot™ Vision Screener (Welch Allyn Inc., Skaneateles Falls, New York, USA). RESULTS: A total of 150 nurses participated in the study (response rate: 30.0%). The mean age was 41.2 ± 8.9 years old. Distance spectacles, reading spectacles and both types of spectacles were used by 37 (24.7%), 32 (21.3%) and 10 (6.7%) nurses, respectively. A total of 58 nurses (38.7%) failed the vision screening test. Visual defects were detected for the first time in 13 nurses (8.7%). With regards to regular eye checkups, 77 participants (51.3%) reported acceptable ocular healthcare-seeking behaviours; this factor was significantly associated with age and the use of visual aids (P <0.01 each). CONCLUSION: A high proportion of participants failed the vision screening tests and only half displayed good ocular healthcare-seeking behaviours. This is concerning as ophthalmic nurses are likely to face fewer barriers to eye care services than the general population.

Competitive Growth Enhances Conditional Growth Mutant Sensitivity to Antibiotics and Exposes a Two-Component System as an Emerging Antibacterial Target in Burkholderia cenocepacia.

Chemogenetic approaches to profile an antibiotic mode of action are based on detecting differential sensitivities of engineered bacterial strains in which the antibacterial target (usually encoded by an essential gene) or an associated process is regulated. We previously developed an essential-gene knockdown mutant library in the multidrug-resistant Burkholderia cenocepacia by transposon delivery of a rhamnose-inducible promoter. In this work, we used Illumina sequencing of multiplex-PCR-amplified transposon junctions to track individual mutants during pooled growth in the presence of antibiotics. We found that competition from nontarget mutants magnified the hypersensitivity of a clone underexpressing gyrB to novobiocin by 8-fold compared with hypersensitivity measured during clonal growth. Additional profiling of various antibiotics against a pilot library representing most categories of essential genes revealed a two-component system with unknown function, which, upon depletion of the response regulator, sensitized B. cenocepacia to novobiocin, ciprofloxacin, tetracycline, chloramphenicol, kanamycin, meropenem, and carbonyl cyanide 3-chlorophenylhydrazone, but not to colistin, hydrogen peroxide, and dimethyl sulfoxide. We named the gene cluster esaSR for enhanced sensitivity to antibiotics sensor and response regulator. Mutational analysis and efflux activity assays revealed that while esaS is not essential and is involved in antibiotic-induced efflux, esaR is an essential gene and regulates efflux independently of antibiotic-mediated induction. Furthermore, microscopic analysis of cells stained with propidium iodide provided evidence that depletion of EsaR has a profound effect on the integrity of cell membranes. In summary, we unraveled a previously uncharacterized two-component system that can be targeted to reduce antibiotic resistance in B. cenocepacia.

Understanding the Pathogenicity of Burkholderia contaminans, an Emerging Pathogen in Cystic Fibrosis.

Several bacterial species from the Burkholderia cepacia complex (Bcc) are feared opportunistic pathogens that lead to debilitating lung infections with a high risk of developing fatal septicemia in cystic fibrosis (CF) patients. However, the pathogenic potential of other Bcc species is yet unknown. To elucidate clinical relevance of Burkholderia contaminans, a species frequently isolated from CF respiratory samples in Ibero-American countries, we aimed to identify its key virulence factors possibly linked with an unfavorable clinical outcome. We performed a genome-wide comparative analysis of two isolates of B. contaminans ST872 from sputum and blood culture of a female CF patient in Argentina. RNA-seq data showed significant changes in expression for quorum sensing-regulated virulence factors and motility and chemotaxis. Furthermore, we detected expression changes in a recently described low-oxygen-activated (lxa) locus which encodes stress-related proteins, and for two clusters responsible for the biosynthesis of antifungal and hemolytic compounds pyrrolnitrin and occidiofungin. Based on phenotypic assays that confirmed changes in motility and in proteolytic, hemolytic and antifungal activities, we were able to distinguish two phenotypes of B. contaminans that coexisted in the host and entered her bloodstream. Whole genome sequencing revealed that the sputum and bloodstream isolates (each representing a distinct phenotype) differed by over 1,400 mutations as a result of a mismatch repair-deficient hypermutable state of the sputum isolate. The inferred lack of purifying selection against nonsynonymous mutations and the high rate of pseudogenization in the derived isolate indicated limited evolutionary pressure during evolution in the nutrient-rich, stable CF sputum environment. The present study is the first to examine the genomic and transcriptomic differences between longitudinal isolates of B. contaminans. Detected activity of a number of putative virulence factors implies a genuine pathogenic nature of this novel Bcc species.

Determination of a broad spectrum of pharmaceuticals and endocrine disruptors in biofilm from a waste water treatment plant-impacted river.

Wastewater treatment plants (WWTPs) are one of the main sources of pharmaceuticals and endocrine disrupting compounds in freshwater ecosystems, and several studies have reported bioaccumulation of these compounds in different organisms in those ecosystems. River biofilms are exceptional indicators of pollution, but very few studies have focused on the accumulation of these emerging contaminants. The objectives of this study were first to develop an efficient analytical methodology for the simultaneous analysis of 44 pharmaceuticals and 13 endocrine disrupting compounds in biofilm, and second, to assess persistence, distribution, and bioaccumulation of these contaminants in natural biofilms inhabiting a WWTP-impacted river. The method is based on pressurized liquid extraction, purification by solid-phase extraction, and analysis by ultra performance liquid chromatography coupled to a mass spectrometer (UPLC-MS/MS) in tandem. Recoveries for pharmaceuticals were 31-137%, and for endocrine disruptors 32-93%. Method detection limits for endocrine disruptors were in the range of 0.2-2.4 ng g(-1), and for pharmaceuticals, 0.07-6.7 ng g(-1). A total of five endocrine disruptors and seven pharmaceuticals were detected in field samples at concentrations up to 100 ng g(-1).

Draft Genome Sequences of Burkholderia contaminans, a Burkholderia cepacia Complex Species That Is Increasingly Recovered from Cystic Fibrosis Patients.

Burkholderia contaminans belongs to the Burkholderia cepacia complex (BCC), a group of bacteria that are ubiquitous in the environment and capable of infecting the immunocompromised and people with cystic fibrosis. We report here draft genome sequences for the B. contaminans type strain LMG 23361 and an Argentinian cystic fibrosis sputum isolate.

An electron transfer flavoprotein is essential for viability and its depletion causes a rod-to-sphere change in Burkholderia cenocepacia.

Essential gene studies often reveal novel essential functions for genes with dispensable homologues in other species. This is the case with the widespread family of electron transfer flavoproteins (ETFs), which are required for the metabolism of specific substrates or for symbiotic nitrogen fixation in some bacteria. Despite these non-essential functions high-throughput screens have identified ETFs as putatively essential in several species. In this study, we constructed a conditional expression mutant of one of the ETFs in Burkholderia cenocepacia, and demonstrated that its expression is essential for growth on both complex media and a variety of single-carbon sources. We further demonstrated that the two subunits EtfA and EtfB interact with each other, and that cells depleted of ETF are non-viable and lack redox potential. These cells also transition from the short rods characteristic of Burkholderia cenocepacia to small spheres independently of MreB. The putative membrane partner ETF dehydrogenase also induced the same rod-to-sphere change. We propose that the ETF of Burkholderia cenocepacia is a novel antibacterial target.

Development of an extraction and purification method for the determination of multi-class pharmaceuticals and endocrine disruptors in freshwater invertebrates.

Aquatic organisms from freshwater ecosystems impacted by waste water treatment plant (WWTP) effluents are constantly exposed to constant concentrations of pharmaceuticals, endocrine disruptors and related compounds, among other anthropogenic contaminants. Macroinvertebrates inhabiting freshwater ecosystems might be useful bioindicators of exposure to contaminants, since their lives are long enough to bioaccumulate, but at the same time may integrate short-term changes in the environment. However, studies about potential bioaccumulation of emerging contaminants in these organisms are very scarce. The objectives of this study were to develop an analytical methodology for the analysis of 41 pharmaceuticals and 21 endocrine disruptors in freshwater invertebrates. In addition, bioaccumulation of these contaminants in three macroinvertebrate taxa inhabiting a waste water treatment plant -impacted river was evaluated. The method for the simultaneous extraction of both families of compounds is based on sonication, purification via removal of phospholipids, and analysis by ultra performance liquid chromatography coupled to a mass spectrometer (UPLC-MS/MS) in tandem. Recoveries for pharmaceuticals were 34-125%, and for endocrine disruptors were 48-117%. Method detection limits (MDLs) for EDCs were in the range of 0.080-2.4 ng g(-1), and for pharmaceuticals, 0.060-4.3 ng g(-1). These pollutants were detected in water samples taken downstream the waste water treatment plant effluent at concentrations up to 572 ng L(-1). Two non-esteroidal anti-inflammatory drugs, diclofenac and ibuprofen, and four endocrine disruptors - estrone, bisphenol A, TBEP, and nonylphenol - were detected in at least one macroinvertebrate taxa in concentrations up to 183 ng g(-1) (dry weight). An isobaric interference was identified during the analysis of diclofenac in Hydropsyche samples, which was successfully discriminated via accurate mass determination by TFC-LTQ Orbitrap.

Burkholderia cenocepacia conditional growth mutant library created by random promoter replacement of essential genes.

Identification of essential genes by construction of conditional knockouts with inducible promoters allows the identification of essential genes and creation of conditional growth (CG) mutants that are then available as genetic tools for further studies. We used large-scale transposon delivery of the rhamnose-inducible promoter, PrhaB followed by robotic screening of rhamnose-dependent growth to construct a genomic library of 106 Burkholderia cenocepacia CG mutants. Transposon insertions were found where PrhaB was in the same orientation of widely conserved, well-characterized essential genes as well as genes with no previous records of essentiality in other microorganisms. Using previously reported global gene-expression analyses, we demonstrate that PrhaB can achieve the wide dynamic range of expression levels required for essential genes when the promoter is delivered randomly and mutants with rhamnose-dependent growth are selected. We also show specific detection of the target of an antibiotic, novobiocin, by enhanced sensitivity of the corresponding CG mutant (PrhaB controlling gyrB expression) within the library. Modulation of gene expression to achieve 30-60% of wild-type growth created conditions for specific hypersensitivity demonstrating the value of the CG mutant library for chemogenomic experiments. In summary, CG mutants can be obtained on a large scale by random delivery of a tightly regulated inducible promoter into the bacterial chromosome followed by a simple screening for the CG phenotype, without previous information on gene essentiality.

Phenylalanine induces Burkholderia cenocepacia phenylacetic acid catabolism through degradation to phenylacetyl-CoA in synthetic cystic fibrosis sputum medium.

Synthetic cystic fibrosis sputum medium (SCFM) is rich in amino acids and supports robust growth of Burkholderia cenocepacia, a member of the Burkholderia cepacia complex (Bcc). Previous work demonstrated that B. cenocepacia phenylacetic acid (PA) catabolic genes are up-regulated during growth in SCFM and are required for full virulence in a Caenorhabditis elegans host model. In this work, we investigated the role of phenylalanine, one of the aromatic amino acids present in SCFM, as an inducer of the PA catabolic pathway. Phenylalanine degradation intermediates were used as sole carbon sources for growth and gene reporter experiments. In addition to phenylalanine and PA, phenylethylamine, phenylpyruvate, and 2-phenylacetamide were usable as sole carbon sources by wild type B. cenocepacia K56-2, but not by a PA catabolism-defective mutant. EMSA analysis showed that the binding of PaaR, the negative regulator protein of B. cenocepacia PA catabolism, to PA regulatory DNA could only be relieved by phenylacetyl-Coenzyme A (PA-CoA), but not by any of the putative phenylalanine degradation intermediates. Taken together, our results show that in B. cenocepacia, phenylalanine is catabolized to PA and induces PA catabolism through PA activation to PA-CoA. Thus, PaaR shares the same inducer with PaaX, the regulator of PA catabolism in Escherichia coli, despite belonging to a different protein family.

Regulation of phenylacetic acid degradation genes of Burkholderia cenocepacia K56-2.

BACKGROUND: Metabolically versatile soil bacteria Burkholderia cepacia complex (Bcc) have emerged as opportunistic pathogens, especially of cystic fibrosis (CF). Previously, we initiated the characterization of the phenylacetic acid (PA) degradation pathway in B. cenocepacia, a member of the Bcc, and demonstrated the necessity of a functional PA catabolic pathway for full virulence in Caenorhabditis elegans. In this study, we aimed to characterize regulatory elements and nutritional requirements that control the PA catabolic genes in B. cenocepacia K56-2. RESULTS: Translational fusions of the PA degradation gene promoters with eGFP were constructed and introduced in B. cenocepacia K56-2. eGFP expression was observed when the reporter strains were grown in minimal media containing glycerol and PA or other compounds expected to proceed through the PA pathway, and in synthetic CF medium (SCFM). Addition of succinate or glucose to the PA containing medium repressed eGFP expression. To show that BCAL0210, a putative TetR-type regulator gene encodes a regulator for the PA genes in B. cenocepacia, we developed a BCAL0210 insertional mutant reporter strain. Results show that these strains exhibit fluorescence regardless of the presence of PA in the culture. CONCLUSION: The PA catabolic genes of B. cenocepacia K56-2 are induced by PA and other related compounds, are negatively regulated by PaaR (named herein), a TetR-type regulator, and are subjected to catabolic repression by glucose and succinate. As the PA catabolic pathway of B. cenocepacia appears to be induced during growth in synthetic cystic fibrosis medium (SCFM), further research is necessary to determine the relevance of this pathway in CF-like conditions and in other host-pathogen interactions.

Human soluble tumor necrosis factor receptor I (sTNF-RI) and interleukin-I receptor antagonist (IL-I Ra) in different stages of acute rheumatic fever.

OBJECTIVE: Acute rheumatic fever (ARF) results from an autoimmune response to infection with group A streptococci. Serum concentrations of two anti-inflammatory cytokines, interleukin-I receptor antagonist (IL-IRa) and human soluble tumor necrosis factor receptor I (sTNF-RI) were determined in patients with ARF at the time of admission and 3 months after treatment in order to evaluate changes in cytokine concentrations occurring during different stages of the disease. METHODS: Serum concentrations of two anti-inflammatory cytokines, IL-I Ra and sTNF-RI , were investigated in children with ARF at the time of admission (n=21) and after 3 months following the cessation of treatment (n=15). The sTNF-RI and sIL-IRa were measured quantitatively in serum using enzyme-linked immunosorbent assay (ELISA). RESULTS: Levels of IL-1Ra and sTNF-RI were found to be significantly higher during acute phase and remission period of ARF when compared to age-matched healthy controls (p=0.001 and p=0.0001, respectively). CONCLUSION: Our study demonstrated that two anti-inflammatory cytokines, serum sTNFRI and IL-1Ra, are increased in acute and remission stages of ARF reflecting activation of the cellular immune response. We suggest this increase might probably be generated in an effort to counteract the already increased concentrations of proinflammatory cytokines.

Inverse relationship between the ratio of ICAM-1 expressing lymphocytes and serum TGF-beta 1 concentrations in acute rheumatic fever.

Autoimmunity in acute rheumatic fever (ARF) is triggered by group-A beta hemolytic streptococci (GAS). Although most of the recent work has focused on the major impact of lymphocytes, the exact immunopathogenesis is still unresolved. Regulation of self-tolerance in response to GAS has been investigated in various animal experiments. This study aimed to associate the ratio of lymphocytes bearing adhesion/costimulatory molecules, Bcl-2/CD95 and serum TGF-beta1 concentrations with clinical stages of ARF. Thirty-five patients were assigned according to the clinical stages. Bcl-2 expression on CD19+ and CD3+ lymphocytes was similar within patient groups and controls. CD62p expression was higher in patients with carditis. The ratio of ICAM-1 bearing lymphocytes was significantly different between patient groups and controls and was increased through acute to remission stages longitudinally. In contrast, a gradual and significant decrease in TGF-beta1 concentrations was observed longitudinally from acute to chronic stages. A positive correlation has been documented between ICAM-1+ lymphocyte ratios and Fas+ cytotoxic T cell ratios supported by a prominent increase in CD95+ T cells. These data draw our attention to the role of ICAM-1, Fas and TGF-beta1 in ARF pathogenesis through the perspective of self-tolerance in a clinical setting.