No abstract

This study was undertaken to provide a comprehensive set of data relevant to disclosing the physiological effects and possible oxygen transport limitations in the Chinook salmon (Oncorhynchus tshawytscha) during an acute temperature change. Fish were instrumented with a blood flow probe around the ventral aorta and catheters in the dorsal aorta and sinus venosus. Water temperature was progressively increased from 13 degrees C in steps of 4 degrees C up to 25 degrees C. Cardiac output increased from 29 to 56 ml.min(-1).kg(-1) between 13 and 25 degrees C through an increase in heart rate (58 to 105 beats/min). Systemic vascular resistance was reduced, causing a stable dorsal aortic blood pressure, yet central venous blood pressure increased significantly at 25 degrees C. Oxygen consumption rate increased from 3.4 to 8.7 mg.min(-1).kg(-1) during the temperature increase, although there were signs of anaerobic respiration at 25 degrees C in the form of increased blood lactate and decreased pH. Arterial oxygen partial pressure was maintained during the heat stress, although venous oxygen partial pressure (Pv(O(2))) and venous oxygen content were significantly reduced. Cardiac arrhythmias were prominent in three of the largest fish (>4 kg) at 25 degrees C. Given the switch to anaerobic metabolism and the observation of cardiac arrhythmias at 25 degrees C, we propose that the cascade of venous oxygen depletion results in a threshold value for Pv(O(2)) of around 1 kPa. At this point, the oxygen supply to systemic and cardiac tissues is compromised, such that the oxygen-deprived and acidotic myocardium becomes arrhythmic, and blood perfusion through the gills and to the tissues becomes compromised.

Solving the antibiotic resistance crisis requires the discovery of new antimicrobial drugs and the preservation of existing ones. The discovery of inhibitors of antibiotic resistance, antibiotic adjuvants, is a proven example of the latter. A major difficulty in identifying new antibiotics is the frequent rediscovery of known compounds, necessitating laborious "dereplication" to identify novel chemical entities. We have developed an antibiotic resistance platform (ARP) that can be used for both the identification of antibiotic adjuvants and for antibiotic dereplication. The ARP is a cell-based array of mechanistically distinct individual resistance elements in an identical genetic background. In dereplication mode, we demonstrate the rapid identification, and thus discrimination, of common antibiotics. In adjuvant discovery mode, we show that the ARP can be harnessed in screens to identify inhibitors of resistance. The ARP is therefore a powerful tool that has broad application in confronting the resistance crisis.

Plazomicin is a next-generation, semisynthetic aminoglycoside antibiotic currently under development for the treatment of infections due to multidrug-resistant Enterobacteriaceae. The compound was designed by chemical modification of the natural product sisomicin to provide protection from common aminoglycoside modifying enzymes that chemically alter these drugs via N-acetylation, O-adenylylation, or O-phosphorylation. In this study, plazomicin was profiled against a panel of isogenic strains of Escherichia coli individually expressing twenty-one aminoglycoside resistance enzymes. Plazomicin retained antibacterial activity against 15 of the 17 modifying enzyme-expressing strains tested. Expression of only two of the modifying enzymes, aac(2')-Ia and aph(2″)-IVa, decreased plazomicin potency. On the other hand, expression of 16S rRNA ribosomal methyltransferases results in a complete lack of plazomicin potency. In vitro enzymatic assessment confirmed that AAC(2')-Ia and APH(2'')-IVa (aminoglycoside acetyltransferase, AAC; aminoglycoside phosphotransferase, APH) were able to utilize plazomicin as a substrate. AAC(2')-Ia and APH(2'')-IVa are limited in their distribution to Providencia stuartii and Enterococci, respectively. These data demonstrate that plazomicin is not modified by a broad spectrum of common aminoglycoside modifying enzymes including those commonly found in Enterobacteriaceae. However, plazomicin is inactive in the presence of 16S rRNA ribosomal methyltransferases, which should be monitored in future surveillance programs.

Resistance to the antibiotic fusidic acid (FA) in the human pathogen Staphylococcus aureus usually results from expression of FusBtype proteins (FusB or FusC). These proteins bind to elongation factor G (EF-G), the target of FA, and rescue translation from FAmediated inhibition by an unknown mechanism. Here we show that the FusB family are two-domain metalloproteins, the C-terminal domain of which contains a four-cysteine zinc finger with a unique structural fold. This domain mediates a high-affinity interaction with the C-terminal domains of EF-G. By binding to EF-G on the ribosome, FusB-type proteins promote the dissociation of stalled ribosome·EF-G·GDP complexes that form in the presence of FA, thereby allowing the ribosomes to resume translation. Ribosome clearance by these proteins represents a highly unusual antibiotic resistance mechanism, which appears to be fine-tuned by the relative abundance of FusB-type protein, ribosomes, and EF-G.antibacterial | protein synthesis | translocation | structure