Resistance genomics and molecular epidemiology of high-risk clones of ESBL-producing Pseudomonas aeruginosa in young children.

Introduction: The emergence of multidrug-resistant Pseudomonas aeruginosa poses a global threat, but the distribution and resistance profiling are unclear, especially in young children. Infections due to P. aeruginosa are common, associated with high mortality, and increasingly ß-lactam drug resistant. Methods: We studied the molecular epidemiology and antibiotic resistance mechanisms in 294 clinicalisolates of P. aeruginosa from a pediatric hospital in China. Non-duplicate isolates were recovered from clinical cases and were identified using an API-20 kit followed by antimicrobial susceptibility testing using the VITEK®2 compact system (BioMerieux, France) and also by broth dilution method. In addition, a double-disc synergy test for the ESBL/E-test for MBL was performed. The presence of beta-lactamases, plasmid types, and sequence types was determined by PCR and sequencing. Results: Fifty-six percent (n = 164) of the isolates were resistant to piperacillin-tazobactam, followed by cefepime (40%; n = 117), ceftazidime (39%; n = 115), imipenem (36%; n = 106), meropenem (33%; n = 97), and ciprofloxacin (32%; n = 94). Forty-two percent (n = 126) of the isolates were positive for ESBL according to the double-disc synergy test. The blaCTX-M-15 cephalosporinase was observed in 32% (n = 40/126), while 26% (n = 33/126) werepositive for blaNDM-1 carbapenemase. Aminoglycoside resistance gene aac(3)IIIawas observed in 16% (n = 20/126), and glycylcyclines resistance gene tet(A) was observed in 12% (n = 15/126) of the isolates. A total of 23 sequence types were detected, including ST1963 (12%; n = 16), followed by ST381 (11%; n = 14), ST234 (10%; n = 13), ST145 (58%; n = 10), ST304 (57%; n = 9), ST663 (5%; n = 7), and a novel strain. In ESBL-producing P. aeruginosa, 12 different Incompatibility groups (Inc) were observed, the most common being IncFI, IncFIS, and IncA/C. The MOBP was the most common plasmid type, followed by MOBH, MOBF, and MOBQ. Discussion: Our data suggest that the spread of antibiotic resistance is likely due toclonal spread and dissemination of different clinical strains of P. aeruginosa harbouring different plasmids. This is a growing threat in hospitals particularly in young children which needs robust prevention strategies.

Perfusion MR prior to radiotherapy is a strong predictor of survival in high-grade gliomas after proton and carbon ion radiotherapy.

Background: To assess the survival predictability of perfusion magnetic resonance imaging (MRI) by the normalized cerebral blood volume (nCBV) prior to particle beam radiotherapy (PBRT) in high-grade glioma (HGG) patients underwent particle therapy. Methods: The study retrieved dynamic susceptibility contrast MRI acquired prior to PBRT between 6/2015 and 3/2019 in 45 patients with HGG. Maximum nCBV (nCBVmax) within or adjacent to surgical/tumor bed was measured using 'hot-spot' method. The predictive values of nCBVmax for progression-free survival (PFS) and overall survival (OS) were assessed in univariate Kaplan-Meier curve and multivariate Cox proportional hazards (CPH) models. Nomograms based on CPH results were constructed to individualize the predicted probability of OS and PFS. Results: The Kaplan-Meier curves and all CPH models based on nCBVmax as continuous variable (nCBVmax-C), group by cut-off derived from median value and Youden-index method showed that nCBVmax prior to radiotherapy was a strong predictor for both PFS and OS in HGG patients who underwent PBRT. Nomograms built on CPH models showed similar excellent performance in both discrimination and calibration. Conclusions: Perfusion imaging prior to PBRT is a strong predictor of survival in HGG. Novel perfusion MR-based nomogram with prospective validation could potentially be formally used in future clinical practice to individualize survival probability.

Particle beam radiation therapy for head and neck rhabdomyosarcoma in adults.

Background: Rhabdomyosarcoma (RMS) is rare in adults, with a significantly worse prognosis than its pediatric counterpart. Radiotherapy (RT) plays a significant role in treating head and neck RMS (HNRMS), but the outcomes of conventional RT are limited by the complex anatomy and unfavorable pathology subtypes of the adult H&N RMS. Here, we aim to report the effectiveness and safety of carbon-ion beam RT (CIRT), either alone or in combination with proton radiotherapy (PRT) in the management of adult HNRMS. Methods: Fifteen adult patients with HNRMS were enrolled on a prospective registry protocol between 06/2015 and 12/2019. Eight patients presented with parameningeal tumors, and eight had unfavorable pathology subtypes [alveolar =7, not otherwise specified (NOS) =1]. Eleven patients had gross tumors before the start of RT (volume range, 46.1-137.6 cm3). Two patients failed the earlier RT. All except for one patient received multi-drug chemotherapy. The median absolute dose of particle beam RT was 70.0 Gy [relative biological effectiveness (RBE)]. Results: With a median follow-up of 21 months, local or distant recurrence occurred in three and four patients, respectively, and two added patients had both local and distant failure. One patient died of distant metastasis (DM), and another died of an unrelated condition. The 1- and 2-year overall survival (OS), local relapse-free survival (LRFS), distant metastasis-free survival (DMFS), and progression-free survival (PFS) rates were 87.5% and 70.0%, 92.3% and 67.1%, 72.2% and 54.2%, and 65.0% and 24.4%, respectively, for the entire cohort. Both patients who failed earlier RT and received salvage CIRT developed DM but were alive at last follow-up. No acute toxicity of ≥ grade 3 or late toxicity of ≥ grade 2 was observed. Conclusions: CIRT, either used alone or in combination with PRT, is not only feasible and safe but also useful in local disease control for HNRMS. DM is the most important cause of treatment failure; thus, more effective systemic treatment is needed to improve the prognosis of HNRMS further.

A cross-sectional analysis of registered clinical trials on the use of particle beam radiation therapy in head and neck cancers.

Background: To present and analyze the current status of registered clinical trials on particle beam (including proton and carbon ion beam) radiation therapy (PBRT) for head and neck (H&N) malignancies, and to provide insights for future clinical research, we designed the cross-sectional analysis. Methods: We identified and analyzed all clinical trials of interest registered on ClinicalTrials.gov and PTCOG.ch until March 22, 2020. Results: We identified 57 registered clinical trials related to the use of proton therapy or carbon ion radiation therapy (CIRT) in H&N malignancies. There were 20, 27, and 5 trials focused on CIRT, proton therapy, and both ions, respectively. The eligible trials were registered between 2007 and 2020, mainly focused on adenoid cystic carcinoma (ACC), squamous cell carcinoma (SCC), sinonasal malignancies (SNM), skull base tumors, locally advanced, and recurrent tumors. The nature of 23 (40%) trials were not stated and could not be identified. A total of 25 (44%) registered trials were phase II, including randomized controlled trials (RCTs). There were 14 RCTs (7 phase II, 2 phase II/III, 2 phase III, 1 phase I/II, and 2 phase not applicable), and 25 studies including RCTs were registered before the first enrolment. There were 11 completed clinical trials among the eligible trials, including 7 with published trial-related results. Conclusions: Less than 10% of the countries with PBRT treatment facilities in operation have initiated clinical trials on H&N cancer. Furthermore, among all registered trials, less than 10% have been completed with results published. More clinical trials, especially high quality trials, are needed for optimizing and standardizing treatment techniques of PBRT for H&N malignancies.

Prevalence, Antibiotic-Resistance, and Replicon-Typing of Salmonella Strains among Serovars Mainly Isolated from Food Chain in Marche Region, Italy.

Nontyphoidal salmonellosis (NTS) is the second most commonly reported gastrointestinal infection in humans and an important cause of food-borne outbreaks in Europe. The use of antimicrobial agents for animals, plants, and food production contributes to the development of antibiotic-resistant Salmonella strains that are transmissible to humans through food. The aim of this study was to investigate the presence and the potential dissemination of multidrug-resistant (MDR) Salmonella strains isolated in the Marche Region (Central Italy) via the food chain. Strains were isolated from different sources: food, human, food animal/livestock, and the food-processing environment. Among them, we selected MDR strains to perform their further characterization in terms of resistance to tetracycline agent, carriage of tet genes, and plasmid profiles. Tetracycline resistance genes were detected by PCR and plasmid replicons by PCR-based replicon typing (PBRT). A total of 102 MDR Salmonella strains were selected among the most prevalent serovars: S. Infantis (n = 36/102), S. Derby (n = 20/102), S. Typhimurium (n = 18/102), and a monophasic variant of S. Typhimurium (MVST, n = 28/102). Resistance to sulfisoxazole (86%) and tetracycline (81%) were the most common, followed by ampicillin (76%). FIIS was the most predominant replicon (17%), followed by FII (11%) and FIB (11%) belonging to the IncF incompatibility group. Concerning the characterization of tet genes, tetB was the most frequently detected (27/89), followed by tetA (10/89), tetG (5/89), and tetM (1/89). This study showed the potential risk associated with the MDR Salmonella strains circulating along the food chain. Hence, epidemiological surveillance supported by molecular typing could be a very useful tool to prevent transmission of resistant Salmonella from food to humans, in line with the One Health approach.

The Co-occurrence of mcr-3 and fosA3 in IncP plasmid in ST131 Escherichia coli: A novel case.

INTRODUCTION: Plasmid-mediated colistin resistance genes, especially mcr-3 combined with the fosfomycin resistance gene fosA3, are a grave health concern. Our study was designed to determine the epidemiological characteristics of the combination of mcr-3 and fosA3 in Anhui province, China. METHODOLOGY: A total of 127 multi-drug-resistant (MDR) E. coli strains were assessed for antibiotic resistance/sensitivity to detect mcr-3 and fosA3 using polymerase chain reaction (PCR) and sequencing. The genes of interest were conjugated using EC600, and replicon and sequence types (STs) were identified by PCR-based replicon typing (PBRT) and multilocus sequence typing (MLST). Cluster similarity and genomic relatedness among the positive isolates were confirmed by Xbal PFGE. RESULTS: The processed E. coli isolates were highly resistant to the tested antibiotics; the prevalence of mcr-3 was 0.78% in the transferable IncP-type plasmid in ST131, whereas fosA3 prevalence was 38.58% among different transferable plasmids, including IncFIIK, IncFII and IncA/C, and in various STs including ST69, ST1193, ST12, ST46, ST57, ST1196, ST38, ST95, ST131, ST7584 and ST10184. Both were successfully transferred to EC600. The Xbal PFGE cluster exposed similarities among the STs. CONCLUSIONS: Our results show that to control the spread of colistin and fosfomycin resistance genes in human pathogens, the ban on colistin must be continued in animal feeding farms not only in China but around the world; additionally, awareness platforms on the use of colistin must be implemented and strict policies in poultry and pig farms must be maintained. Furthermore, fosfomycin misuse by patients and overuse by physicians must be strictly managed to stop the spread of fosfomycin resistance.

Validity and reliability of 6-minute pegboard and ring test in patients with asthma.

OBJECTIVE: The 6-minute pegboard and ring test (6PBRT) is a test of upper-extremity functional capacity designed for and validated in chronic obstructive pulmonary disease. The aim of this study was to evaluate the validity and reliability of the 6PBRT in asthma patients. METHODS: Thirty-four adults (30 women, 4 men) with well-controlled asthma were included. Unsupported upper-extremity exercise capacity was assessed using 6PBRT, maximal arm exercise capacity using an arm ergometer, handgrip strength using a hand dynamometer, activities of daily living with the London Chest Activities of Daily Living Scale (LCADL), Milliken ADL scale (MAS) and health-related quality of life using the Asthma Quality of Life Questionnaire (AQLQ) and Health Assessment Questionnaire Disability Index (HAQ-DI). RESULTS: The 6PBRT showed moderate to excellent test-retest reliability with an intraclass correlation coefficient (ICC) value of 0.872 [95% confidence interval (CI) 0.702-0.941]. The 6PBRT was reproducible according to Bland-Altman analysis, with upper and lower limits of agreement of 53.51 and -25.08 rings moved, respectively. The 6PBRT score was significantly correlated with maximum workload (r = 0.514, p = 0.002) achieved in the arm ergometer test, change in dyspnea during 6PBRT (r = -0.402, p = 0.020), LCADL-self-care (r = -0.364, p = 0.037), MAS total (r = 0.483, p = 0.005), AQLQ-symptom domain (r = 0.420, p = 0.026) and HAQ-DI total scores (r = -0.390, p = 0.025). CONCLUSIONS: The 6PBRT can be used as a valid and reliable test to evaluate functional arm exercise capacity in patients with well-controlled asthma.

Determination of incompatibility group plasmids and copy number of the bla NDM-1 gene in carbapenem-resistant Klebsiella pneumoniae strains recovered from different hospitals in Kerman, Iran.

Introduction. New Delhi metallo-ß-lactamase (NDM)-producing Klebsiella pneumoniae has become a serious global health concern.Hypothesis/Gap Statement. Due to the high genetic diversity among NDM-positive K. pneumoniae, we need further surveillance and studies to better understand the relationships between them. In addition, the coexistence of several plasmid replicon types in NDM-positive K. pneumoniae may affect the copy number of bla NDM, the MIC level to antibiotics, as well as increasing the chance of horizontal gene transfer.Aim. The aim of this study was to determine incompatible plasmid groups and copy numbers of bla NDM, and to investigate the genetic relationship of 37 NDM-positive K. pneumoniae in Kerman, Iran.Methodology. The bla NDM-1 gene was detected and confirmed by PCR-sequencing. The plasmid replicon types were determined by PCR-based replicon typing (PBRT) and the copy number of bla NDM-1 was determined by quantitaive real time-PCR (qPCR). Random amplified polymorphic DNA (RAPD)-PCR typing was used to detect genetic relationships between the strains.Results. In this study, 10 different replicon types, including Frep [n=25 (67.5 %)], FIIAs [n=11 (29.7 %)], FIA [n=5 (13.5 %)], FIB [n=3 (8.1 %)], I1-Iγ [n=2 (5.4 %)], L/M [n=7 (18.9 %)], A/C [n=7 (18.9 %)], Y [n=3 (8.1 %)], P [n=1 (2.7 %)] and FIC [n=1 (2.7 %)] were reported. The copy numbers of the bla NDM-1 gene varied from 30.00 to 5.0×106 and no statistically significant correlation was observed between a rise of the MIC to imipenem and the copy numbers of bla NDM-1 (P>0.05). According to RAPD typing results, 35 strains were divided into five clusters, while two strains were non-typeable.Conclusion. The spread of NDM-1-producing K. pneumoniae strains that carry several plasmid replicon types increases the chance of horizontal transfer of antibiotic resistance genes in hospital settings. In this study, 10 different replicon types were identified. We could not find any relationship between the increase of MIC levels to imipenem and the copy numbers of bla NDM-1. Therefore, due to the identification of different replicon types in this study, the type and genetic characteristics of bla NDM-1-carrying plasmids, and other factors such as antibiotic selective pressure, probably affect the copy number of bla NDM-1 and change the MIC level to imipenem.

Plasmid Replicon Typing of Antibiotic-Resistant Escherichia coli From Clams and Marine Sediments.

Unlike human isolates, environmental Escherichia coli isolates have not been thoroughly investigated for the diversity and transferability of antibiotic-resistant plasmids. In this study, antibiotic-resistant strains from marine sediment (n = 50) and clams (n = 53) were analyzed (i) for their plasmid content using a PCR-based plasmid replicon typing (PBRT) kit and (ii) for the transferability of plasmid-associated antibiotic resistance (AR) traits by mating experiments. Fifteen of the thirty replicons targeted by the PBRT kit were detected in the isolates; 8/15 were identified in both sediment and clam isolates, although at different frequencies. The most frequent replicons in sediment (74%) and in clam strains (66%) alike, were FIA, FIB, or FII, which are associated with the IncF group, followed by the I1α replicon, which was more frequent in clam (24.5%) than in sediment (10%) strains. More than 50% of the strains contained multiple replicons; although 15 were untypable, S1-PFGE analysis demonstrated that 14/15 carried no plasmids. All cryptic strains were successfully typed and were positive for IncF or IncI replicons. Antibiotic-resistant strains accounted for 63% of all isolates and were significantly (p < 0.05) more frequent in phylogroup A. Most (35%) multidrug-resistant (MDR) strains belonged to phylogroup A, too. Although 25/26 MDR strains were positive for IncF plasmids (the exception being a clam strain), the FII-FIB rep combination was predominant (63%) among the sediment isolates, whereas most clam isolates (40%) carried the FII replicon alone. In mating experiments, selected MDR strains carrying FIB, FII, and I1α replicons, used as the donors, transferred multiple ARs together with the IncF or IncI plasmids at high frequency. Since IncI plasmids are common in E. coli and Salmonella enterica isolates from poultry, our findings suggest an animal origin to the E. coli clam strains carrying IncI plasmids. They also suggest a role for IncI plasmids in the spread of ARs among environmental Enterobacteriaceae and, through the food chain, to human isolates. In conclusion, the PBRT kit proved to be a useful tool to identify plasmids carrying antibiotic-resistant genes and to shed light on the factors underpinning their diffusion.

Emergence of ß-lactamase- and carbapenemase- producing Enterobacteriaceae at integrated fish farms.

BACKGROUND: Epidemiological studies suggested that determinants for antibiotic resistance have originated in aquaculture. Recently, the integrated agriculture-aquaculture system has been implemented, where fish are raised in ponds that receive agriculture drainage water. The present study aims to investigate the occurrence of ß-lactamase and carbapenemase-producing Enterobacteriaceae in the integrated agriculture-aquaculture and the consequent public health implication. METHODS: Samples were collected from fish, fishpond water inlets, tap water, outlet water, and workers at sites of integrated agriculture-aquacultures. Samples were also taken from inhabitants of the aquaculture surrounding areas. All samples were cultured on MacConkey agar, the Enterobacteriaceae isolates were tested for susceptibility to cephalosporins and carbapenems, and screened for blaCTX-M-15, blaSHV, blaOXA-1, blaTEM, blaPER-1, blaKPC, blaOXA-48, and blaNDM. Strains having similar resistance phenotype and genotype were examined for the presence of Incompatible (Inc) plasmids. RESULTS: A major proportion of the Enterobacteriaceae isolates were resistant to cephalosporins and carbapenems. Among the 66 isolates from fish, 34 were resistant to both cephalosporin and carbapenem groups, 26 to carbapenems alone, and 4 to cephalosporins alone. Of the 15 isolates from fishpond water inlets, 8 showed resistance to both groups, 1 to carbapenems alone, and 5 to cephalosporins alone. Out of the 33 isolates from tap water, 17 were resistant to both groups, and 16 to cephalosporins alone. Similarly, of the 16 outlet water isolates, 10 were resistant to both groups, and 6 to cephalosporins alone. Furthermore, of the 30 examined workers, 15 carried Enterobacteriaceae resistant strains, 10 to both groups, and 5 to cephalosporins alone. Similar strains were isolated from the inhabitants of the aquaculture surrounding areas. Irrespective of source of samples, strains resistant to all examined antibiotics, carried predominantly the carbapenemase gene blaKPC either alone or with the ß-lactamase genes (blaCTX-M-15, blaSHV, blaTEM, and blaPER-1). The isolates from fish, water, and workers harboured a wide-range of multi-drug-resistance Inc. plasmids, which were similar among all isolates. CONCLUSION: The present findings suggest transmission of the resistance genes among Enterobacteriaceae strains from different sources. This reiterates the need for control strategies that focus on humans, animals, water, and sewage systems to solve the antibiotic resistance problem.

Changes in electricity production and microbial community evolution in constructed wetland-microbial fuel cell exposed to wastewater containing Pb(II).

Two constructed wetland microbial fuel cell (CW-MFC) devices, experimental group (EG, with 5 mg/L Pb(II) addition) and control group (CG) were built to explore the changes in power generation, wastewater purification and microbial community structure under Pb(II) stress. The voltage of EG (343.16 ± 12.14 mV) was significantly higher (p < 0.01) than that of CG (295.49 ± 13.91 mV), and the highest power density of the EG and CG were 7.432 mW·m-2 and 3.873 mW·m-2, respectively. There was no significant difference in the removal of common pollutants between these groups except for the NH4+-N removal efficiency, which was probably caused by the inhibition of the bioactivity of Comamonas (AOB) in the anode of the experimental group by Pb(II). Pb(II) was effectively removed by CW-MFC (84.86 ± 3%), and the abundant amount of fulvic acid-like matter in the extracellular polymeric substance (EPS) of the EG contributed to its removal. The presence of Pb(II) had a negative effect on both microbial community diversity and species richness. The abundance of a lead resistance gene, pbrT, decreased with long-term Pb(II) pressure. This is evidence of microbial adaptation to Pb(II).

Plasmid Typing and Classification.

Plasmids identification and classification is an essential parameter in current bacterial typing. The most widely used PCR-based methods are the PCR-based replicon typing (PBRT) and the degenerate primer MOB typing (DPMT). PBRT targets the replicons on the plasmids and DPMT targets the relaxase genes. A finer resolution of phylogenetic relatedness can be obtained by plasmid multiLocus sequence typing available for the major plasmid types occurring in Enterobacteriaceae.

The Inc FII Plasmid and its Contribution in the Transmission of blaNDM-1 and blaKPC-2 in Klebsiella pneumoniae in Egypt.

The emergence of blaKPC-2 and blaNDM-1 producing Klebsiella pneumoniae represents a great problem in many Egyptian hospitals. One hundred and twenty-six K. pneumoniae isolates from patients admitted to Assiut University Hospital were identified by an API20E kit. Carbapenemase-producing K. pneumoniae (CPKP) was detected by the modified carbapenem inactivation method (mCIM), the EDTA-modified carbapenem inactivation method (eCIM), and an E-test. Based on the polymerase chain reaction, all isolates were negative for bla-VIM-1 and bla-IMP-1, fifteen of these isolates were positive for both blaKPC-2 and blaNDM-1, two isolates were positive for blaKPC-2 only, and twenty-eight isolates were positive for bla-NDM-1 only. Although one isolate was positive for the string test, all CPKP isolates were negative for capsular genes. Only 71.1% of CPKP transferred their plasmids to their corresponding transconjugants (E. coli J53). The resistance patterns of the clinical isolates and their transconjugates were similar, except for 12 isolates, which showed differences with their transconjugates in the resistance profile of four antibiotics. Molecular typing of the plasmids based on replicon typing showed that Inc FIIK and FII plasmids predominated in isolates and their transconjugants carrying blaKPC-2 and/or blaNDM-1. Conjugative Inc FII plasmids play an important role in the spread of CPKP, and their recognition is essential to limit their spread.

Evolution and resistance of a microbial community exposed to Pb(II) wastewater.

This study investigated the treatment performance of activated sludge on Pb(II)-containing wastewater, including contaminant removal efficiency, extracellular polymeric substances, pbrT gene content and the microbial community. The average removal efficiencies of ammonia nitrogen, chemical oxygen demand, total phosphorus, total nitrogen and Pb(II) were 40% ±â€¯4%, 91% ±â€¯3%, 95% ±â€¯3%, 51% ±â€¯5% and 92% ±â€¯9% during the stable operation stage, respectively. Moreover, the extracellular polymeric substance -protein contents increased significantly from day 0 to day 60 (p < 0.05). The most abundant fluorescent component in extracellular polymeric substances was a humic acid-like substance, and its fluorescence intensity increased significantly from day 0 to day 60 (p < 0.05). Adsorption of negatively charged organic functional groups in extracellular polymeric substances was identified as a major component of the removal of Pb(II). Most of the denitrifying bacteria associated with nitrogen removal showed an increasing trend during the acclimation stage, which may have resulted in high total nitrogen removal efficiency. In addition, pbrT uptake protein was found to be responsible for the uptake of Pb(II) into cells. The abundance of the pbrT gene showed a downward trend (p < 0.05) after adding Pb(II), probably because expression of the pbrT gene was inhibited under Pb(II) stress. Sphingopyxis containing the pbrT gene was the dominant resistance genus, and its relative abundance increased significantly (p < 0.05) from day 0 to day 60. This study provided a theoretical basis for the treatment of Pb(II)-containing wastewater.

Characterization and Bio-Typing of Multidrug Resistance Plasmids From Uropathogenic Escherichia coli Isolated From Clinical Setting.

Urinary tract infection is primarily caused by Escherichia coli. Multidrug resistance and their rapid dissemination in this pathogenic microbe complicate therapeutic strategies and threaten public health. Conjugation systems responsible for interbacterial transmission of antibiotic resistance are plasmid-encoded and can be classified as the P, F, and I types. Specific pili types and pili associated proteins were related to the transfer among this gram-negative organism and were thought to depend on contacts created by these structures at the time of DNA transport. In this study, conjugation system types of the plasmids that harbor multidrug resistant genes (aac-1b-cr, oqxAB, qnrB, qnrS, bla TEM, bla OXA) amongst 19 E. coli uropathogenic isolates were characterized under ciprofloxacin/ceftazidime selection individually by pili and pili associated gene types. Investigations indicated incidence of single plasmid of multiple replicon type amongst the transconjugants. bla TEM, bla CTX-M, bla OXA, aac-1b-cr, oqxAB, qnrB, qnrS genes in varied combination were observed to be successfully co-transmitted against ceftazidme/ciprofloxacin selection. Seven primer pair sets were selected that encodes pili and pili associated genes (traF, trwJ, traE, trhE, traG, pilM, pilx4) by nucleotide database search tools using annotated plasmids of different incompatibility types to assign the conjugation system type of the transmissible resistant plasmids by PCR. traF was predominant irrespective of drug selection that indicated F-type conjugation system was responsible for transmission of resistant plasmids which results in the rapid dissemination of antibiotic resistance in the isolates screened. Therefore this is a first report of its kind that investigated pili and pili associated genes to bio-type multidrug resistant plasmids and their transmission in clinical settings amongst uropathogenic E. coli circulated in the eastern part of India.

A Simulation Study Using Terrestrial LiDAR Point Cloud Data to Quantify Spectral Variability of a Broad-Leaved Forest Canopy.

In this analysis, a method for construction of forest canopy three-dimensional (3D) models from terrestrial LiDAR was used for assessing the influence of structural changes on reflectance for an even-aged forest in Belgium. The necessary data were extracted by the developed method, as well as it was registered the adjacent point-clouds, and the canopy elements were classified. Based on a voxelized approach, leaf area index (LAI) and the vertical distribution of leaf area density (LAD) of the forest canopy were derived. Canopy⁻radiation interactions were simulated in a ray tracing environment, giving suitable illumination properties and optical attributes of the different canopy elements. Canopy structure was modified in terms of LAI and LAD for hyperspectral measurements. It was found that the effect of a 10% increase in LAI on NIR reflectance can be equal to change caused by translating 50% of leaf area from top to lower layers. As presented, changes in structure did affect vegetation indices associated with LAI and chlorophyll content. Overall, the work demonstrated the ability of terrestrial LiDAR for detailed canopy assessments and revealed the high complexity of the relationship between vertical LAD and reflectance.

High diversity of genes and plasmids encoding resistance to third-generation cephalosporins and quinolones in clinical Escherichia coli from commercial poultry flocks in Italy.

The aim was to investigate occurrence and diversity of plasmid-mediated resistance to third-generation cephalosporins (3GC) and quinolones in clinical Escherichia coli from 200 industrial poultry farms across Italy. E. coli was isolated from colibacillosis lesions in turkeys (n = 109), broilers (n = 98) and layers (n = 22) between 2008 and 2012. 3GC-resistant isolates were screened for extended-spectrum and AmpC ß-lactamase (ESBL/AmpC), while all isolates were tested for plasmid-mediated quinolone resistance (PMQR) genes. ESBL/AmpC- and PMQR-positive isolates were typed by pulsed-field gel electrophoresis and antimicrobial susceptibility testing, and their plasmids were characterised by replicon typing, multilocus sequence typing, restriction fragment length polymorphism and conjugation. EBSL/AmpC genes (blaCTX-M-1, blaCTX-M-14, blaCTX-M-2, blaSHV-12 and blaCMY-2) were detected in 7%, 9% and 4% of isolates from turkeys, broilers and layers, respectively. We identified seven ESBL/AmpC-encoding plasmid types, usually conjugative (78%), with a marked prevalence of IncI1/pST3 plasmids carrying blaCTX-M-1. PMQR occurred less frequently among isolates from turkeys (0.9%) compared to those from broilers (5%) and layers (4%). The PMQR genes qnrS, qnrB19 and oqxA/B were located on three plasmid types and two non-typeable plasmids, mostly (85%) conjugative. ESBL/AmpC- and PMQR-positive isolates were genetically unrelated and 64% of them were additionally resistant to aminoglycosides, sulfonamides and tetracyclines. Our data show that 3GC- and quinolone-resistant clinical E. coli in Italian poultry production represent a highly diverse population often resistant to most antimicrobials available for poultry. These findings underline the crucial need to develop new strategies for prevention and control of colibacillosis.

Diversity of plasmids harboring blaCMY-2 in multidrug-resistant Escherichia coli isolated from poultry in Brazil.

Multidrug-resistance (MDR) has been increasingly reported in Gram-negative bacteria from the intestinal microbiota, environment and food-producing animals. Resistance plasmids able to harbor different transposable elements are capable to mobilize antimicrobial resistance genes and transfer to other bacterial hosts. Plasmids carrying blaCMY are frequently associated with MDR. The present study assessed the presence of plasmid-encoded ampC genes (blacmy, blamox, blafox, blalat, blaact, blamir, bladha, blamor) in commensal E. coli isolated from apparently healthy broiler chickens. Furthermore, we characterized the plasmids and identified those harboring the resistance genes. We isolated 144/200 (72%) of E. coli isolates with resistance to cefotaxime and the resistance gene identified was blaCMY-2. The pulsed-field gel electrophoresis (PFGE) analysis showed high diversity of the genetic profiles. The phylogenetic groups A, B1, B2, and D were identified among E. coli isolates and group D was the most prevalent. The PCR-based replicon typing (PBRT) analysis identified four distinct plasmid incompatibility groups (Inc) in MDR isolates. Moreover, plasmids harboring blaCMY-2, ranged in size from 50kb to 150kb and 51/144 (35%) belonged to IncK, 21/144 (14.5%) to IncB/O, 8/144 (5.5%) to IncA/C, 1/144 (0.5%) to IncI, while 63/144 (44.5%) were not typeable by PBRT. Overall, a high prevalence of blaCMY-2 genes was found in a diverse population of commensal MDR E. coli from poultry in Brazil, harbored into different plasmids.

Metastatic clival chordoma: a case report of multiple extraneural metastases following resection and proton beam radiotherapy in a 5-year old boy.

The authors report the case of a 5-year-old boy in whom extraneural metastases developed 5 years after he underwent an occipitocervical fusion and transoral approach to treat a clival chordoma without local recurrence. Following primary resection, the patient's postoperative course was complicated by recurrent meningitis secondary to CSF leak, which responded to antibiotics, and communicating hydrocephalus, for which a ventriculoperitoneal shunt was placed. The patient then underwent postoperative proton beam radiotherapy. Five years following his initial presentation, surveillance imaging revealed a new asymptomatic lung mass for which the patient underwent thoracotomy and resection of the mass. Histological examination of the lung mass revealed findings consistent with a de-differentiated chordoma, confirming extraneural metastasis from the original tumor without evidence of local recurrence. Chest wall and scalp metastases subsequently developed, and the patient was started on an adjuvant chemotherapy regimen that included imatinib and rapamycin followed by subsequent nivolumab and an EZH2 inhibitor for recurrent, disseminated disease. Despite this patient's remote and distant metastases, primary gross-total resection for chordoma remains a critical treatment objective, followed by proton beam radiotherapy. This case illustrates the importance of interval posttreatment imaging and the emerging potential to treat chordoma with molecularly targeted therapies.

Outcomes following attempted en bloc resection of cervical chordomas in the C-1 and C-2 region versus the subaxial region: a multiinstitutional experience.

OBJECT: Chordomas involving the mobile spine are ideally managed via en bloc resection with reconstruction to optimize local control and possibly offer cure. In the cervical spine, local anatomy poses unique challenges, limiting the feasibility of aggressive resection. The authors present a multi-institutional series of 16 cases of cervical chordomas removed en bloc. Particular attention was paid to clinical outcome, complications, and recurrence. In addition, outcomes were assessed according to position of tumor at the C1-2 level versus the subaxial (SA) spine (C3-7). METHODS: The authors reviewed cases involving patients who underwent en bloc resection of cervical chordoma at 4 large spine centers. Patients were included if the lesion epicenter involved the C-1 to C-7 vertebral bodies. Demographic data and details of surgery, follow-up course, exposure to adjuvant therapy, and complications were obtained. Outcome was correlated with presence of tumor in C1-2 versus subaxial spine via a Student t-test. RESULTS: Sixteen patients were identified (mean age at presentation 55 ± 14 years). Seven cases (44%) cases involved C1-2, and 16 involved the subaxial spine. Median survival did not differ significantly different between the C1-2 (72 months) and SA (60 months) groups (p = 0.65). A combined (staged anteroposterior) approach was used in 81% of the cases. Use of the combined approach was significantly more common in treatment of subaxial than C1-2 tumors (100% vs 57%, p = 0.04). En bloc resection was attempted via an anterior approach in 6% of cases (C1-2: 14.3%; SA: 0%; p = 0.17) and a posterior approach in 13% of cases (C1-2: 29%; SA: 0%; p = 0.09). The most commonly reported margin classification was marginal (56% of cases), followed by violated (25%) and wide (19%). En bloc excision of subaxial tumors was significantly more likely to result in marginal margins than excision of C1-2 tumors (C1-2: 29%; SA: 78%; p = 0.03). C1-2 tumors were associated with significantly higher rates of postoperative complications (C1-2: 71%; SA: 22%; p = 0.03). Both local and distant tumor recurrence was greatest for C1-2 tumors (local C1-2: 29%; local SA: 11%; distant C1-2: 14%; distant SA: 0%). Statistical analysis of tumor recurrence based on tumor location was not possible due to the small number of cases. There was no between-groups difference in exposure to postoperative adjuvant radiotherapy. There was no difference in median survival between groups receiving proton beam radiotherapy or intensity-modulated radiotherapy versus no radiation therapy (p = 0.8). CONCLUSIONS: Compared with en bloc resection of chordomas involving the subaxial cervical spine, en bloc resection of chordomas involving the upper cervical spine (C1-2) is associated with poorer outcomes, such as less favorable margins, higher rates of complications, and increased tumor recurrence. Data from this cohort do not support a statistically significant difference in survival for patients with C1-2 versus subaxial disease, but larger studies are needed to further study survival differences.