Presence of antibiotic-resistant commensal bacteria in samples from agricultural, city, and national park environments evaluated by standard culture and real-time PCR methods

This study examined the presence of antibiotic-resistant commensal bacteria among cattle operations representing areas heavily affected by agriculture, city locations representing areas affected by urban activities and indirectly affected by agriculture, and a national park representing an area not affected by agriculture. A total of 288 soil, fecal floor, and water samples were collected from cattle operations, from the city of Fort Collins, and from Rocky Mountain National Park (RMNP) in Colorado. In addition, a total of 42 new and unused feed, unused bedding, compost, and manure samples were obtained from the cattle operations. Total, tetracycline-resistant, and ceftiofur-resistant bacterial populations were enumerated by both standard culture plating and real-time PCR methods. Only wastewater samples from the cattle operations demonstrated both higher tetracycline-resistant bacterial counts (enumerated by the culture plating method) and tetracycline resistance gene copies (quantified by real-time PCR) compared to water samples collected from non-farm environments. The ceftiofur resistance gene, blaCMY-2, was not detectable in any of the samples, while the tetracycline resistance genes examined in this study, tet(B), tet(C), tet(W), and tet(O), were detected in all types of tested samples, except soil samples from RMNP. Tetracycline resistance gene pools quantified from the tet(O) and tet(W) genes were bigger than those from the tet(B) and tet(C) genes in fecal and water samples. Although only limited resistance genes, instead of a full set, were selected for real-time PCR quantification in this study, our results point to the need for further studies to determine natural and urban impacts on antibiotic resistance.     

Complete genome sequence of Rhizobium leguminosarum bv. trifolii strain WSM1325, an effective microsymbiont of annual Mediterranean clovers

Rhizobium leguminosarum bv trifolii is a soil-inhabiting bacterium that has the capacity to be an effective nitrogen fixing microsymbiont of a diverse range of annual Trifolium (clover) species. Strain WSM1325 is an aerobic, motile, non-spore forming, Gram-negative rod isolated from root nodules collected in 1993 from the Greek Island of Serifos. WSM1325 is produced commercially in Australia as an inoculant for a broad range of annual clovers of Mediterranean origin due to its superior attributes of saprophytic competence, nitrogen fixation and acid-tolerance. Here we describe the basic features of this organism, together with the complete genome sequence, and annotation. This is the first completed genome sequence for a microsymbiont of annual clovers. We reveal that its genome size is 7,418,122 bp encoding 7,232 protein-coding genes and 61 RNA-only encoding genes. This multipartite genome contains 6 distinct replicons; a chromosome of size 4,767,043 bp and 5 plasmids of size 828,924 bp, 660,973 bp, 516,088 bp, 350,312 bp and 294,782 bp.     

Osmoregulatory fluid intake but not hypovolemic thirst is intact in mice lacking angiotensin

Water intakes in response to hypertonic, hypovolemic, and dehydrational stimuli were investigated in mice lacking angiotensin II as a result of deletion of the angiotensinogen gene (Agt-/- mice), and in C57BL6 wild-type (WT) mice. Baseline daily water intake in Agt-/- mice was approximately threefold that of WT mice because of a renal developmental disorder of the urinary concentrating mechanisms in Agt-/- mice. Intraperitoneal injection of hypertonic saline (0.4 and 0.8 mol/l NaCl) caused a similar dose-dependent increase in water intake in both Agt-/- and WT mice during the hour following injection. As well, Agt-/- mice drank appropriate volumes of water following water deprivation for 7 h. However, Agt-/- mice did not increase water or 0.3 mol/l NaCl intake in the 8 h following administration of a hypovolemic stimulus (30% polyethylene glycol sc), whereas WT mice increased intakes of both solutions during this time. Osmoregulatory regions of the brain [hypothalamic paraventricular and supraoptic nuclei, median preoptic nucleus, organum vasculosum of the lamina terminalis (OVLT), and subfornical organ] showed an increased number of neurons exhibiting Fos-immunoreactivity in response to intraperitoneal hypertonic NaCl in both Agt-/- mice and WT mice. Polyethylene glycol treatment increased Fos-immunoreactivity in the subfornical organ, OVLT, and supraoptic nuclei in WT mice but only increased Fos-immunoreactivity in the supraoptic nucleus in Agt-/- mice. These data show that brain angiotensin is not essential for the adequate functioning of neural pathways mediating osmoregulatory thirst. However, angiotensin II of either peripheral or central origin is probably necessary for thirst and salt appetite that results from hypovolemia.     

The effect of a low-protein diet in pregnancy on offspring renal calcium handling

Low birth weight humans and rats exposed to a low-protein diet in utero have reduced bone mineral content. Renal calcium loss during the period of rapid skeletal growth is associated with bone loss. Because young rats exposed to low protein display altered renal function, we tested the hypothesis that renal calcium excretion is perturbed in this model. Pregnant Wistar rats were fed isocalorific diets containing either 18% (control) or 9% (low) protein throughout gestation. Using standard renal clearance techniques, Western blotting for renal calcium transport proteins, and assays for Na(+)-K(+)-ATPase activity and serum calcitropic hormones, we characterized calcium handling in 4-wk-old male offspring. Histomorphometric analyses of femurs revealed a reduction in trabecular bone mass in low-protein rats. Renal calcium (control vs. low protein: 10.4 +/- 2.1 vs. 27.6 +/- 4.5 nmol x min(-1) x 100 g body wt(-1); P < 0.01) and sodium excretion were increased, but glomerular filtration rate was reduced in low-protein animals. Total plasma calcium was reduced in low-protein rats (P < 0.01), but ionized calcium, serum calcitropic hormone concentrations, and total body calcium did not differ. There was no significant change in plasma membrane Ca(2+)-ATPase pump, epithelial calcium channel, or calbindin-D(28K) expression in low-protein rat kidneys. However, Na(+)-K(+)-ATPase activity was 36% lower (P < 0.05) in low-protein rats. These data suggest that the hypercalciuria of low-protein rats arises through a reduction in passive calcium reabsorption in the proximal tubule rather than active distal tubule uptake. This may contribute to the reduction in bone mass observed in this model.     

Spatio-temporal distribution patterns and conservation of fish assemblages in a Chilean coastal river

River environments are characterized by extreme spatial and temporal variation in the physical environment. The relationship of fish assemblages to environmental variation is poorly understood in many systems. In Chile zonation patterns of fish assemblages have been documented in several Andean river drainages. Coastal river drainages are comparatively small, but inordinately important because of their highly endemic flora and fauna and their proximity to major human populations. For conservation purposes it is important to understand what environmental factors affect assemblage structure of fishes especially in the comparatively high diversity coastal drainages. We studied patterns of fish distribution and abundance in three rivers of the coastal, Andalien drainage near Concepción, Chile. We used multi-dimensional scaling analyses to compare patterns among zones (rithron, transition and potamon) and high and low flow seasons. Species assemblages differed by zone, but not with season. Assemblages consisted of nested subsets of species characterized by their range of distribution among zones. One species group was composed of widespread species that occurred in all three zones, another species group consisted of species found only in transitional and potamal zones, and a final group was comprised of species found only in the potamal zone. The potamal zone contained the most diverse and abundant fish assemblage. Fish assemblages were related to both water quality and habitat structure variables. This study suggests that the key to conserving the diversity of native fish communities in coastal Chilean rivers is in the conservation of potamal regions. Unfortunately, most protected areas in Chile are in the depauperate headwaters of drainages. Protection of only headwaters is clearly inadequate and will not contribute to the conservation of this unique freshwater fish fauna.     

MCH-R1 antagonists based on an arginine scaffold: SAR studies on the amino-terminus

We have identified a novel series of potent MCH-R1 antagonists based on l-arginine. As predicted by computational methods, there was an activity dependence on the pi-electronic character of the aromatic systems corresponding to the amino-terminus of these molecules. These results have enhanced our understanding of the MCH-R1 receptor and the potential for a predictive homology model.     

Mucosal HIV-1 transmission and prevention strategies in BLT humanized mice

Clinical trials testing microbicides and related biomedical interventions to block HIV transmissions have produced contradictory results and to date it is unclear why. Further elucidation of the molecular basis of mucosal HIV transmission and extensive pharmacokinetic and pharmacodynamic analyses are essential to implementing effective prevention strategies. Animal models are of critical importance to this effort and bone marrow-liver-thymus (BLT) humanized mice have recently emerged as a powerful small animal research platform for in vivo efficacy evaluation of mucosal and parenteral HIV-1 prevention interventions. The availability of this validated system for the pre-clinical evaluation of HIV-1 prevention approaches will accelerate the implementation of the best candidate interventions into clinical trials.     

Lower lip suspension using bilateral temporalis muscle flaps and fascia lata grafts

Oral incompetence following composite reconstruction of total and subtotal lower lip defects without any functioning lower lip muscle is a difficult problem for reconstructive surgeons. The authors retrospectively reviewed the use of a novel bilateral temporalis suspension technique for oral incompetence following lower lip reconstruction over a 10-year period. The timing of the reconstruction, cause of the defect, period of follow-up, and any complications were noted. Three cases of lower lip resuspension using bilateral temporalis flaps and fascia lata grafts were performed from 2000 to 2010. Two cases were secondary to burn trauma and one was from ballistic trauma. All patients underwent traditional means of reconstruction using free microvascular composite tissue transfer with and without fascial slings. All three patients presented with persistent lower lip incompetence. The average interval between the initial reconstructive operations and the resuspension operations was 1.6 years. All patients achieved dynamic oral competence at the first postoperative visit. At a mean follow-up of 3.6 years, all patients had maintained lower lip function. Dynamic lower lip resuspension with bilateral temporalis flaps and fascia lata grafts is an option for refractory lower lip drooping following total and subtotal loss, especially after conventional static reconstruction and without any functional orbicularis muscle. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.