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While most detailed analyses of antibiotic resistance plasmids focus on those found in clinical isolates, less is known about the vast environmental reservoir of mobile genetic elements and the resistance and virulence factors they encode. We selectively isolated three strains of cefotaxime-resistant Escherichia coli from a wastewater-impacted coastal wetland. The cefotaxime-resistant phenotype was transmissible to a lab strain of E. coli after one hour, with frequencies as high as 10-3 transconjugants per recipient. Two of the plasmids also transferred cefotaxime resistance to Pseudomonas putida, but these were unable to back-transfer this resistance from P. putida to E. coli. In addition to the cephalosporins, E. coli transconjugants inherited resistance to at least seven distinct classes of antibiotics. Complete nucleotide sequences revealed large IncF-type plasmids with globally distributed replicon sequence types F31:A4:B1 and F18:B1:C4 carrying diverse antibiotic resistance and virulence genes. The plasmids encoded extended-spectrum β-lactamases blaCTX-M-15 or blaCTX-M-55, each associated with the insertion sequence ISEc9, although in different local arrangements. Despite similar resistance profiles, the plasmids shared only one resistance gene in common, the aminoglycoside acetyltransferase aac(3)-IIe. Plasmid accessory cargo also included virulence factors involved in iron acquisition and defense against host immunity. Despite their sequence similarities, several large-scale recombination events were detected, including rearrangements and inversions. In conclusion, selection with a single antibiotic, cefotaxime, yielded conjugative plasmids conferring multiple resistance and virulence factors. Clearly, efforts to limit the spread of antibiotic resistance and virulence among bacteria must include a greater understanding of mobile elements in the natural and human-impacted environments.

Medulloblastoma is among the most common malignant brain tumors in children. Recent studies have identified at least four subgroups of the disease that differ in terms of molecular characteristics and patient outcomes. Despite this heterogeneity, most patients with medulloblastoma receive similar therapies, including surgery, radiation, and intensive chemotherapy. Although these treatments prolong survival, many patients still die from the disease and survivors suffer severe long-term side effects from therapy. We hypothesize that each patient with medulloblastoma is sensitive to different therapies and that tailoring therapy based on the molecular and cellular characteristics of patients' tumors will improve outcomes. To test this, we assembled a panel of orthotopic patient-derived xenografts (PDX) and subjected them to DNA sequencing, gene expression profiling, and high-throughput drug screening. Analysis of DNA sequencing revealed that most medulloblastomas do not have actionable mutations that point to effective therapies. In contrast, gene expression and drug response data provided valuable information about potential therapies for every tumor. For example, drug screening demonstrated that actinomycin D, which is used for treatment of sarcoma but rarely for medulloblastoma, was active against PDXs representing Group 3 medulloblastoma, the most aggressive form of the disease. Functional analysis of tumor cells was successfully used in a clinical setting to identify more treatment options than sequencing alone. These studies suggest that it should be possible to move away from a one-size-fits-all approach and begin to treat each patient with therapies that are effective against their specific tumor. SIGNIFICANCE: These findings show that high-throughput drug screening identifies therapies for medulloblastoma that cannot be predicted by genomic or transcriptomic analysis.

Chelated copper (Cu) sources, such as Cu glycinate (CuGly), may be more bioavailable relative to Cu sulfate (CuSO 4 ) when fed to ruminants under antagonistic pressure. The objective of this study was to determine the bioavailability of CuGly (GemStone Cu; Phibro Animal Health) relative to CuSO4 in steers fed a diet supplemented with 0.3% sulfur and 2 mg molybdenum/kg of dry matter (DM). Sixty Angus crossbred steers (n  = 12 per treatment) averaging 288 ± 4.85 kg were enrolled in a 90-d study and fed a corn silage-based diet with one of five Cu supplementation strategies, including no supplemental Cu (CON), 5 or 10 mg supplemental Cu from CuSO4/kg DM, and 5 or 10 mg supplemental Cu from CuGly/kg DM. Steers were housed in pens equipped with GrowSafe feed bunks (GrowSafe Systems Ltd. Airdire, AB, Canada), with six steers per pen. Growth performance, liver Cu, and plasma Cu were analyzed in the MIXED procedure of SAS 9.4 (SAS Inst. Inc, Cary, NC) with orthogonal contrasts to compare CON vs. 5 mg Cu/kg DM, CON vs. 10 mg Cu/kg DM, 5 vs. 10 mg Cu/kg DM, and CuSO4 vs. CuGly. Copper indices were regressed against Cu intake and slopes were calculated using the GLM procedure SAS. Dietary Cu supplementation did not affect steer body weights on days 0, 28, 56, or 90 (P  ≥ 0.52), average daily gain, dry matter intake, or gain:feed (P  ≥ 0.36). Final plasma Cu concentration did not differ between CON vs. 5 mg Cu/kg DM (P  = 0.79), CON vs. 10 mg Cu/kg DM (P  = 0.65), or 5 vs. 10 mg Cu/kg DM (P  = 0.39). Steers receiving CuSO4 tended to have greater final plasma Cu concentrations than those receiving CuGly (P  = 0.08). Initial liver Cu concentration averaged 374 mg Cu/kg DM, which is considered highly adequate. No steers reached deficient Cu status by the end of the 90-d period. Control steers had lesser final liver Cu concentrations than supplemented steers (P  ≤ 0.04). Steers receiving 10 mg supplemental Cu/kg DM had greater liver Cu concentrations than those receiving 5 mg supplemental Cu/kg DM (P  = 0.01). Copper source had no effect on final liver Cu concentrations (P  = 0.57) and based on liver Cu and Cu intake the bioavailability of CuGly was similar to CuSO4 (115%; P  = 0.27). The initially high Cu status and the fact that cattle did not become Cu deficient may have impacted the relative bioavailability results, and more research is needed to investigate the role initial Cu status and antagonistic pressure play in the bioavailability of chelated Cu sources. [ABSTRACT FROM AUTHOR]

Copper (Cu) from Cu sulfate (CuSO4) may be more susceptible to antagonism when fed in cattle diets high in sulfur and molybdenum, but other Cu supplements may be better able to avoid this. The objective of this study was to determine the bioavailability of Cu carbonate (CuCO3; Emerald-C, Old Bridge Minerals, Old Bridge, NJ) and tribasic Cu chloride (TBCC; Intellibond-C, Selko, Indianapolis, IN) relative to CuSO4 when fed to steers supplemented with 2 mg Mo/kg DM and 0.1% S. Angus crossbred steers [n = 84; body weight (BW) = 282 ± 19.3 kg] were enrolled (n = 12 per treatment) in a 96-d study and fed a corn-silage based diet with one of seven Cu supplementation strategies: no supplemental Cu (CON), 5 or 10 mg supplemental Cu/kg DM from CuSO4, 5 or 10 mg supplemental Cu/kg DM from TBCC, or 5 or 10 mg supplemental Cu/kg DM from CuCO3. Steers were housed in pens equipped with GrowSafe feed bunks (GrowSafe Systems Ltd., Airdire, AB, Canada) with six steers per pen. The GLM procedure of SAS 9.4 was used to determine the bioavailability of CuCO3 and TBCC relative to CuSO4 using both liver and plasma Cu. The MIXED procedure of SAS was used to analyze liver Cu, plasma Cu, and growth performance data. Contrasts compared CON vs. 5 mg/ kg, CON vs. 10 mg/kg, 5 mg/kg vs. 10 mg/kg, CuCO3 vs. CuSO4, TBCC vs. CuSO4, and CuCO3 vs. TBCC. The bioavailability of CuCO3 relative to CuSO4 based on liver and plasma Cu was 89 (P < 0.01) and 133% (P = 0.21), respectively. The bioavailability of TBCC relative to CuSO4 based on liver and plasma Cu was 88 (P < 0.01) and 100% (P = 0.82), respectively. Over the 96-d trial, liver Cu decreased 66% in CON, from 74 to 21 mg Cu/kg DM. Steers receiving CuCO3 had greater plasma Cu concentrations than those receiving TBCC on d 56 (P = 0.05) but were similar by d 96 (P = 0.39). Copper supplemented steers had greater dry matter intake (DMI) than CON (P = 0.01). Steers supplemented with CuCO3 had greater DMI than those receiving TBCC (P = 0.05). Steers receiving TBCC tended to have greater gain:feed than those receiving CuSO4 (P = 0.08), but gain:feed did not differ between CON and supplemented treatments (P ≥ 0.6). In conclusion, CuCO3 and TBCC may be less available to steers under moderate antagonistic pressure based on liver Cu, but Emerald-C CuCO3 may result in greater Cu in circulation. More research is needed to evaluate the impact of feeding CuCO3 on systemic Cu metabolism and how factors such as initial Cu status impact relative bioavailability results. [ABSTRACT FROM AUTHOR]

Angus-cross steers [n = 144; body weight (BW) = 359 kg ± 13.4] were used to assess the effect of dietary Mn and steroidal implants on trace mineral (TM) status, and liver and serum metabolites related to N metabolism. Steers (n = 6/pen) were stratified by BW in a 3 × 2 factorial. GrowSafe bunks recorded individual feed intake (experimental unit = steer; n = 24/ treatment). Dietary treatments included (MANG; 8 pens per treatment; Mn as MnSO4): 1) no supplemental Mn (analyzed 14 mg Mn/kg DM; Mn0); 2) 20 mg supplemental Mn/kg DM (Mn20); 3) 50 mg supplemental Mn/kg DM (Mn50). Within MANG, steers received a steroidal implant treatment (IMP) on d 0: 1) no implant; NO; or 2) combination implant (Revalor-200; REV). Liver biopsies for TM analysis and qPCR, and blood for serum urea-N (SUN) analysis were collected on d 0, 20, 40, and 77. Liver TM, serum metabolite, enzyme activity, and gene expression data were analyzed as repeated measures using PROC MIXED of SAS 9.4. The Correlation Procedure of SAS 9.4 was utilized to assess correlations between liver arginase activity, SUN concentrations, and liver Mn concentrations. No MANG × IMP effects were noted (P ≥ 0.12) for growth or carcass measures. Steroidal implants increased final BW (FBW), overall average daily gain (ADG), and gain to feed (G:F; P ≤ 0.01). Dietary Mn did not influence FBW, overall ADG, or overall G:F (P ≥ 0.14). Liver Mn concentration increased with supplemental Mn (P = 0.01). An IMP × DAY effect was noted for liver Mn (P = 0.01) where NO and REV were similar on d 0 but NO cattle increased liver Mn from d 0 to 20 while REV liver Mn decreased. Relative expression of liver Mn transporter ZnT10 tended to be least (MANG; P = 0.10) in Mn20 while Mn0 and Mn100 were similar. Relative expression of liver Mn transporter ZIP8 was not influenced by MANG, IMP, or MANG × IMP (P ≥ 0.23). Liver arginase activity was not influenced by MANG, IMP, MANG × DAY, IMP × DAY, or MANG × IMP (P ≥ 0.15) but tended to decrease over time (DAY; P = 0.10). On d 0, 20, and 40, arginase activity was correlated (r ≤ 0.33; P ≤ 0.01) with SUN and liver Mn. Relative expression of MnSOD in the liver was greater in REV (P = 0.03) compared with NO and within a MANG × IMP effect (P = 0.01) REV increased liver MnSOD activity. This study indicates current NASEM Mn recommendations are adequate to meet demands of finishing beef cattle given a steroidal implant. This study also reveals how implants and supplemental Mn influence liver enzyme activity and genes related to arginine metabolism, urea synthesis, antioxidant capacity, and TM homeostasis. [ABSTRACT FROM AUTHOR]

The objective of this study was to evaluate the effect of copper (Cu) status on response to lubabegron fumarate (Experior, Elanco, Greenfield, IN). Angus crossbred steers [n = 48; body weight (BW) = 469 ± 31 kg] were enrolled in a 77-d finishing study, housed in pens of 6, with a GrowSafe bunk (GrowSafe Systems, Ltd, Airdire, AB, Canada) in each pen and implanted (Revalor-200, Merck Animal Health, Madison, NJ; 200 mg TBA + 20 mg E2) on d 0. A 2 × 2 factorial arrangement was applied to Cu status (deficient, DEF, no supplemental Cu; or adequate, ADE, 10 mg supplemental Cu/kg DM as CuSO4) and Experior (36 mg·steer-1·d-1, EXP; or no Experior, NOEXP). A prior trial resulted in steers with distinct liver Cu (DEF and ADE were 42 ± 21 and 193 ± 31 mg Cu/kg DM, respectively). Experior was fed from d 37 to d 73, and cattle harvested on d 78. Body weights were collected on d -1, 0, 37, 55 (mid-Experior), 76 and 77. Blood was collected on d 0, 37, 55, and 76 for serum urea nitrogen (SUN), non-esterified fatty acids (NEFA), glucose, and insulin. Data were analyzed in Proc Mixed of SAS as a factorial design, with steer as the experimental unit (n = 12 per treatment). From d 0-37, DEF daily gain and feed efficiency (G:F) were greater than ADE (P < 0.01), while dry matter intake (DMI) did not differ (P = 0.36). Overall DMI was greater in ADE than DEF (P = 0.04) and not affected by Experior (P = 0.13). Carcass-adjusted average daily gain was not affected by Cu or Experior (P ≥ 0.21). Deficient steers had greater carcass-adjusted G:F than ADE (P = 0.02). Hot carcass weight was greater in EXP (P = 0.04), but post-hoc analysis of the Cu x Experior effect (P = 0.2) reveals this was driven by DEF-EXP (15 kg over DEF-NOEXP) rather than ADE-EXP (4 kg over ADE-NOEXP). Marbling tended to be greater in ADE (P = 0.06) and was unaffected by Experior (P = 0.14). The revised quantitative insulin sensitivity check index displayed an EXP x Day effect (P < 0.01) where NOEXP was consistent across days and EXP had greater insulin sensitivity during the Experior period. Serum urea nitrogen was affected by Cu × Experior × Day (P = 0.01), where DEF-NOEXP had lesser d 0 SUN than all others, and EXP decreased on d 55 while NOEXP increased steadily during the finishing phase; by d 76 DEF-EXP remain lesser than NOEXP treatments with ADE-EXP being similar to all others. These data suggest Cu concentration influences steer response to lubabegron fumarate, and more research is needed to determine the cause of this. [ABSTRACT FROM AUTHOR]