bluuebunny/crossref_metadata_2025 huggingface dataset:
Bactericidal killing activities of cefepime, ceftazidime, cefotaxime, and ceftriaxone against Staphylococcus aureus and beta-lactamase-producing strains of Enterobacter aerogenes and Klebsiella pneumoniae in an in vitro infection model
Authors: Palmer S M , Kang S L , Cappelletty D M , Rybak M J
Published: June 2012

Cefepime (CP) is a new injectable cephalosporin with a broad spectrum of activity and stability against common chromosomally and plasmid-mediated beta-lactamases. The bactericidal activities of CP, ceftazidime (CZ), cefotaxime (CTX), and ceftriaxone (CAX) against reference and clinical strains of Staphylococcus aureus, an isogenic pair of Enterobacter aerogenes strains (wild type and a CZ-resistant derepressed mutant), and a Klebsiella pneumoniae isolate possessing a TEM-10 beta-lactamase were investigated in a two-compartment pharmacodynamic in vitro infection model which simulates human pharmacokinetics. An inoculum of approximately 10(6) CFU/ml was used in all model experiments. Antibiotics were administered to simulate the following regimens: CP at 2 g every 12 h (q12h), CZ at 2 g q8h, CTX at 2 g q8h, and CAX at 2 g q24h. Human albumin was added during experiments with CAX and staphylococci to simulate protein binding. Samples were removed at multiple time points over a 48-h period to determine the inoculum size for time-kill curves. Development of resistance was detected by inoculating samples obtained at 0, 24, and 48 h onto antibiotic-containing agar plates. The time to 99.9% killing was used to compare drug regimens. Against staphylococci, the time to bacterial eradication was significantly delayed with CAX-albumin. All regimens had similar activities against the wild-type Enterobacter strain; however, regrowth was noted with CZ, CTX, and CAX against the CZ-resistant strain. There were no differences between the CP, CTX, and CAX regimens against K. pneumoniae. Of interest, no regrowth of any organism was noted with CP. These data indicate that CP has activity against S.aureus and CZ-resistant gram-negative bacilli.
Use of primers selective for vancomycin resistance genes to determine van genotype in enterococci and to study gene organization in VanA isolates
Authors: Miele A , Bandera M , Goldstein B P
Published: June 2012

Vancomycin resistance in enterococci is an emerging therapeutic problem. Resistance is not always detected by standard microbiological methods. Oligonucleotide primers for PCR were designed to target amplification of defined regions of genes of the vanA cluster, as well as vanB and vanC1. These primers correctly identified 30 vancomycin-resistant isolates tested (17 VanA, 7 VanB, and 6 Enterococcus gallinarum). No amplification was observed with Enterococcus casseliflavus or vancomycin-susceptible strains. Using PCR and Southern blotting, we found that all 17 VanA isolates had orf-1, orf-2, vanR, vanS, vanH, vanA, and vanY genes in the same sequence and that the intergenic distances in the vanR-vanA segments were the same. The described methods should be applicable to the rapid detection of the different vancomycin resistance genotypes in enterococci.
Efficacy of D0870 compared with those of itraconazole and amphotericin B in two murine models of invasive aspergillosis
Authors: Denning D W , Hall L , Jackson M , Hollis S
Published: June 2012

D0870 is a novel azole antifungal compound. It was compared with conventional amphotericin B and itraconazole therapy in two murine models of invasive aspergillosis, one a systemic nonimmunocompromised mouse model and the other a temporarily neutropenic mouse respiratory model. D0870 was given orally and achieved measurable concentrations in serum approximately proportional to the daily dose with accumulation over time if it was given twice daily. Amphotericin B at 3.3 mg/kg of body weight was given intraperitoneally for four to six doses, and itraconazole was given orally in a cyclodextrin suspension at 5 to 50 mg/kg daily or twice daily (BID). The duration of therapy varied from 7 to 14 days. In the nonimmunocompromised mouse model, D0870 at 25 mg/kg BID was slightly inferior to amphotericin B and itraconazole with regard to mortality, with a median survival of 20 days for the three groups (P = 0.03 compared with amphotericin B). However, D0870 at 25 mg/kg BID was inferior to amphotericin B (but not itraconazole) with respect to renal culture (P = 0.01) and brain culture (P = 0.0001) results. Only amphotericin B was statistically superior to controls with regard to mortality. In the neutropenic mouse respiratory model, D0870 at 50 mg/kg/day was superior to amphotericin B, itraconazole, and controls with regard to mortality. D0870 at both 25 and 50 mg/kg/day was statistically superior to controls with regard to lung culture results (P = 0.004 to 0.04). A second experiment with a higher inoculum showed that no drug regimen was effective in that model. In all models low doses and concentrations of D0870 in serum were ineffective. D0870 has some efficacy for the treatment of invasive aspergillosis when it is given at modest doses.
Inhibition of growth of Histoplasma capsulatum yeast cells in human macrophages by the iron chelator VUF 8514 and comparison of VUF 8514 with deferoxamine
Authors: Newman S L , Gootee L , Stroobant V , van der Goot H , Boelaert J R
Published: June 2012

Histoplasma capsulatum requires intracellular iron to survive and multiply within human and murine macrophages (M phi). Thus, iron chelators may be useful compounds in the treatment of histoplasmosis. In the present study we compared the efficacies of five different iron chelators with deferoxamine (DEF) for their capacity to inhibit the growth of H. capsulatum yeast cells in culture medium and within human M phi. Of the agents tested, only one, VUF 8514, a 2,2'-bipyridyl analog, was found to be effective. VUF 8514 inhibited the growth of yeast cells in tissue culture medium and within M phi in a dose-response fashion. In tissue culture medium, the 50% effective dose (ED50) of VUF 8514 was 30 nM and the ED50 of DEF was 1 mM. In human M phi, the ED50 of VUF 8514 was 520 nM and the ED50 of DEF was 4 mM. Thus, VUF 8514 was effective at a concentration 7.7 x 10(3)-fold lower than DEF in inhibiting the growth of yeast cells in M phi. Inhibition of the intracellular growth of yeast cells by VUF 8514 was reversed by holotransferrin and iron nitriloacetate, an iron compound that is soluble at neutral to alkaline pH. Thus, VUF 8514 inhibits the intracellular growth of yeast cells by acting as an iron chelator rather than through its capacity as a weak base. These data suggest that the hydroxamic acid siderophore of H. capsulatum yeast cells competes successfully for iron against some iron chelators but not others and that VUF 8514 may be a potential therapeutic agent for the treatment of histoplasmosis.
Pharmacodynamics of metronidazole determined by a time-kill assay for Trichomonas vaginalis
Authors: Nix D E , Tyrrell R , Müller M
Published: June 2012

The pharmacodynamic effects of metronidazole on Trichomonas vaginalis have been poorly characterized. The present in vitro study was performed to characterize the relationship between killing of trichomonads and metronidazole exposure (metronidazole concentration and time of exposure). Five laboratory strains and five recent clinical isolates of T. vaginalis were studied. The minimum lethal concentrations (MLCs) of metronidazole for the strains ranged from 0.8 to 25 micrograms/ml under anaerobic conditions. Metronidazole exhibited concentration-dependent killing against T. vaginalis at concentrations ranging from 0.1 to > 10 times the MLC. The endpoint measurement, the kill rate constant, which was derived from the reduction in the logarithm of the colony count divided by exposure time, compared with the kill rate constant for the growth control was not affected by the time of assessment between 2 and 24 h. The kill rate constant-versus-metronidazole exposure curves were similar when concentration was expressed as a multiple of the MLC. There were no apparent differences between the clinical isolates and laboratory strains. These data suggest that peak metronidazole concentration and/or area under the plasma concentration-versus-time curve are the important pharmacodynamic parameters to be optimized.
Influence of hemodialysis on plasma concentration-time profiles of mefloquine in two patients with end-stage renal disease: a prophylactic drug monitoring study
Authors: Crevoisier C A , Joseph I , Fischer M , Graf H
Published: June 2012

Prophylactic drug monitoring of mefloquine and its carboxylic acid metabolite were studied in two patients with end-stage renal disease undergoing long-term hemodialysis treatment. The patients, short-term travellers to areas where malaria is endemic, took 250 mg of mefloquine (Lariam) once weekly for 2 weeks before and during their 3-week stay abroad and for one week after their return. Pre- and postdialysis blood samples were drawn before their departure and after their return. The concentration-time profiles of mefloquine and its metabolite in plasma samples taken before and after the 3- to 4-h dialysis sessions were similar. Mefloquine and its metabolite could not be detected in the dialysate. These findings show that mefloquine and its metabolite are not, or are very poorly, removed by hemodialysis. Concentrations in plasma and accumulation kinetics were similar to those reported for healthy volunteers and were associated with high prophylactic efficacy against malaria. No special dosage adjustments have to be made in patients undergoing hemodialysis treatment to achieve concentrations in plasma similar to those in healthy volunteers. The prophylactic dose of mefloquine could be given before, during, or after the hemodialysis session.
Effects of zinc-desferrioxamine on Plasmodium falciparum in culture
Authors: Chevion M , Chuang L , Golenser J
Published: June 2012

The zinc-desferrioxamine (Zn-DFO) complex is considered to be more permeative into parasitized erythrocytes than is the metal-free DFO. The former may penetrate the cell and exchange its bound zinc for ferric ions, rendering the iron unavailable for vital parasite functions. The effects of these compounds on the in vitro development of Plasmodium falciparum are compared. The results indicate that Zn-DFO is superior to DFO, especially at concentrations below 20 microM, as shown by decreased levels of hypoxanthine incorporation, lower levels of parasitemia, and interference with the life cycle of the parasite. At low concentrations, DFO even enhanced parasite growth. Such an enhancement was not observed following exposure to Zn-DFO. Experiments in which the compounds were removed from the cultures indicated that parasites treated with Zn-DFO are less likely to recover at a later stage. Since DFO has already been used in humans for the treatment of malaria, its complex with zinc, which is more effective in vitro, should also be examined in vivo.
SB 205952, a novel semisynthetic monic acid analog with at least two modes of action
Authors: Wilson J M , Oliva B , Cassels R , O'Hanlon P J , Chopra I
Published: June 2012

The biological properties of SB 205952, a nitrofuryl oxazole derivative of monic acid, differ from those of the closely related antibacterial agent mupirocin. Compared with mupirocin, SB 205952 has increased antimicrobial potency, an extended spectrum including mupirocin-resistant staphylococci, and rapid bactericidal activity. SB 205952, like mupirocin, is a potent inhibitor of bacterial isoleucyl-tRNA synthetase (IRS) in mupirocin-susceptible organisms but does not inhibit IRS from mupirocin-resistant staphylococci, indicating that SB 205952 has more than one mechanism of action. SB 205952 rapidly inhibits protein, RNA, and DNA syntheses in mupirocin-susceptible and mupirocin-resistant staphylococci. In each case, the effect on RNA synthesis is relaxed by treatment with chloramphenicol, indicating that inhibition of RNA synthesis is probably a secondary consequence of stringent control. It is proposed that SB 205952 possesses one or more mechanisms of action in addition to IRS inhibition, probably mediated by its nitrofuryl component.
Sterol compositions and susceptibilities to amphotericin B of environmental Cryptococcus neoformans isolates are changed by murine passage
Authors: Currie B , Sanati H , Ibrahim A S , Edwards J E , Casadevall A , Ghannoum M A
Published: June 2012

Previous studies have shown that sequential isolates from patients with persistent Cryptococcus neoformans meningoencephalitis can vary in sterol composition and susceptibility to antifungal drugs. To investigate the potential of host factors as mediators of this phenomenon, we compared fungal susceptibilities of environmental and clinical isolates from a limited geographic area. Clinical isolates were less susceptible to amphotericin B than environmental isolates. Five environmental isolates were passaged through BALB/c murine hosts; the passaged isolates had changes in sterol composition and reduced amphotericin B susceptibilities relative to those of the parent isolates. In contrast, murine passage of these isolates did not alter their susceptibilities to fluconazole. The results confirm that changes in sterol composition and antifungal susceptibility can occur in vivo as a result of host factors and suggest that human infection can result in selection of variants with reduced susceptibilities to amphotericin B.
Amphotericin B liposomes with prolonged circulation in blood: in vitro antifungal activity, toxicity, and efficacy in systemic candidiasis in leukopenic mice
Authors: van Etten E W , ten Kate M T , Stearne L E , Bakker-Woudenberg I A
Published: June 2012

Pegylated amphotericin B (AmB) liposomes (PEG-AmB-LIP) were compared with laboratory-prepared nonpegylated AmB liposomes (AmB-LIP), a formulation with a lipid composition the same as that in AmBisome, as well as with industrially prepared AmBisome regarding their in vitro antifungal activities, toxicities, blood residence times, and therapeutic efficacies. Killing of Candida albicans (> 99.9%) during short-term (6-h) incubation was observed at 0.2 mg of AmB per liter for AmB desoxycholate, 0.4 mg of AmB per liter for PEG-AmB-LIP, 0.8 mg of AmB per liter for AmB-LIP, and 12.8 mg of AmB per liter for AmBisome. The maximum tolerated doses of PEG-AmB-LIP, AmB-LIP, and AmBisome were 15, 19, and > 31 mg of AmB per kg of body weight, respectively. In contrast to AmB-LIP, the blood residence time of PEG-AmB-LIP was prolonged and dose independent. In a model of systemic candidiasis in leukopenic mice at a dose of 5 mg of AmB per kg, PEG-AmB-LIP was completely effective and AmB-LIP was partially effective, whereas AmBisome was not effective. AmB-LIP at 11 mg of AmB per kg was partially effective. AmBisome at 29 mg of AmB per kg was completely effective. In conclusion, the therapeutic efficacies of AmB liposomes can be improved by preparing AmB liposomes in which a substantial reduction in toxicity is achieved while antifungal activity is retained. In addition, therapeutic efficacy is favored by a prolonged residence time of AmB liposomes in blood.
Effect of levofloxacin on glycosaminoglycan and DNA synthesis of cultured rabbit chondrocytes at concentrations inducing cartilage lesions in vivo
Authors: Kato M , Takada S , Ogawara S , Takayama S
Published: June 2012

We investigated the toxic effect of levofloxacin (LVFX), a quinolone antibacterial agent, on cartilage by examining aspects of its in vivo toxicokinetics and effect on the function of cultured chondrocytes of the femoral articular cartilage from juvenile New Zealand White rabbits. Repeated administration of LVFX (100 mg/kg) orally for 7 days induced focal necrosis and superficial erosion in the articular cartilage of the femoral condyle, but 30 mg/kg did not. Concentrations of LVFX in the cartilage were highest at the first sampling point (30 min) after a single administration, being 4.93 and 12.2 micrograms/g in the 30- and 100-mg/kg groups, respectively. The arthropathic concentration of LVFX in the cartilage was then shown to be 12.2 micrograms/g or more. For an in vitro study, chondrocytes were separated from the articular cartilage of the rabbit femoral condyle and cultured for 7 days until confluence. 35SO4 uptake by cultured chondrocyte sheets was most susceptible to LVFX, decreasing at drug concentrations of 5 micrograms/ml or more in 24- and 48-h cultures but not in a 72-h culture. Furthermore, 3H-thymidine uptake was decreased at concentrations of 10 micrograms/ml or more in a 48-h culture but not in 24- and 72-h cultures. Rhodamine 123 accumulation was susceptible to inhibition in cultured chondrocytes at an LVFX concentration of 10 micrograms/ml or more. These results suggest that LVFX inhibits glycosaminoglycan synthesis initially and DNA synthesis and mitochondrial function secondarily at actual arthropathic concentrations in cultured rabbit chondrocytes but that these changes are reversible and not enough to kill the cells.
Bactericidal activity against cephalosporin-resistant Streptococcus pneumoniae in cerebrospinal fluid of children with acute bacterial meningitis
Authors: Klugman K P , Friedland I R , Bradley J S
Published: June 2012

There are reports of failure of extended-spectrum cephalosporin treatment in pneumococcal meningitis. On the basis of in vitro and animal experimental studies, the addition of vancomycin or rifampin to an extended-spectrum cephalosporin has been recommended for empiric treatment of these patients. Cerebrospinal fluid (CSF) was taken from 31 children with bacterial meningitis randomized to receive ceftriaxone alone (n = 11), ceftriaxone plus rifampin (n = 10), or ceftriaxone plus vancomycin (n = 10). The CSF from children receiving ceftriaxone alone was unable to kill intermediately ceftriaxone-resistant or fully resistant strains when the concentration of ceftriaxone in the CSF was less than 5 micrograms/ml. At higher concentrations bactericidal activity was present. We have shown that vancomycin penetrates reliably into the CSF of children with acute meningitis, which is in contrast to previous studies with adults. The addition of vancomycin or rifampin to ceftriaxone resulted in significantly enhanced CSF bactericidal activity compared with that of ceftriaxone alone against these resistant strains. Our data suggest that the addition of rifampin or vancomycin to ceftriaxone may be useful for the treatment of cephalosporin-resistant pneumococcal meningitis.
Magnesium deficiency induces joint cartilage lesions in juvenile rats which are identical to quinolone-induced arthropathy
Authors: Stahlmann R , Förster C , Shakibaei M , Vormann J , Günther T , Merker H J
Published: June 2012

Quinolones accumulate in cartilage, and because they form chelate complexes with divalent cations, they possess the potential to induce a deficiency of functionally available magnesium. To test the hypothesis that quinolone-induced arthropathy is caused (or aggravated) by magnesium deficiency in cartilage, we induced magnesium deficiency by feeding juvenile rats a magnesium-deficient diet for 9 days and treated the rats with single oral doses of ofloxacin (0, 100, 300, 600, or 1,200 mg/kg of body weight) during this period. Additional groups of juvenile rats on a normal diet were treated with ofloxacin correspondingly. Typical cartilage lesions (e.g., swollen matrix, cleft formation) were found in knee joints of all magnesium-deficient rats, including those without ofloxacin treatment. Lesions in these groups were not distinguishable from lesions induced by a single dose of 600 mg of ofloxacin per kg of body weight or higher in rats on a normal diet. Ofloxacin levels in plasma after 600 mg/kg of body weight were approximately 10-fold higher than those in humans during therapy with this quinolone. Lesions in rats treated with ofloxacin plus magnesium deficiency were more pronounced than those in rats with normal magnesium concentrations. After intake of a magnesium-deficient diet for 9 days, the magnesium concentration in serum (mean +/- standard deviation) was 0.18 +/- 0.05 mmol/liter (control on normal diet, 0.82 +/- 0.10 mmol/liter). Magnesium concentrations in bone (femur) and cartilage (processus xiphoideus) samples were 64.7 +/- 10.5 and 14.3 +/- 3.9 mmol/kg of dry weight, respectively, which corresponded to approximately 50% of the concentrations measured in controls on a normal diet. It was concluded that quinolone-induced arthropathy is probably caused by a deficit of available magnesium in joint cartilage due to the formation of quinolone-magnesium chelate complexes. If juvenile patients must be treated with quinolones for serious infections, it seems prudent to ensure that these patients do not have a disturbed magnesium balance.
Reduced amoxicillin uptake into human gastric mucosa when gastric juice pH is high
Authors: Cardaci G , Lambert J R , King R G , Onishi N , Midolo P
Published: June 2012

Amoxicillin when administered with gastric acid suppressors has been shown to be effective in eradication of Helicobacter pylori in 50 to 80% of subjects. The aim of this investigator-blind crossover study was to determine if gastric mucosal amoxicillin uptake was affected by increasing gastric juice pH. Fifteen male subjects (7 H. pylori positive and 8 H. pylori negative) were randomized to receive 150 mg of ranitidine twice a day, 300 mg of ranitidine twice a day, or no drug for 2 days prior to upper endoscopy. The last dose of ranitidine was given 60 min prior to upper endoscopy, and amoxicillin (500 mg) was given 30 min prior to upper endoscopy. The amoxicillin concentrations in mucosal biopsy samples, gastric juice, and serum were determined by a standard microbiological bioassay technique. Mean amoxicillin levels were greater in samples of antrum, fundus, and duodenum for volunteers who received no ranitidine than in those receiving 300 mg of ranitidine (P < 0.05) and those receiving 150 mg of ranitidine (P < 0.05 except for fundus). Amoxicillin levels in the antrum, fundus, and duodenum were negatively correlated with gastric juice pH (P < 0.005 for antrum; P < 0.001 for fundus and duodenum). There was no correlation between gastric juice pH and amoxicillin levels in either gastric juice or serum. The amoxicillin concentration in gastric juice was significantly higher with 300 mg of ranitidine than with no ranitidine (P < 0.05). Thus, lower gastric juice pH is associated with a higher rate of mucosal uptake of amoxicillin.
Formulation and efficacy of liposome-encapsulated antibiotics for therapy of intracellular Mycobacterium avium infection
Authors: Oh Y K , Nix D E , Straubinger R M
Published: June 2012

Mycobacterium avium is an intracellular pathogen that can invade and multiply within macrophages of the reticuloendothelial system. Current therapy is not highly effective. Particulate drug carriers that are targeted to the reticuloendothelial system may provide a means to deliver antibiotics more efficiently to M. avium-infected cells. We investigated the formulation of the antibiotics ciprofloxacin and azithromycin in liposomes and tested their antibacterial activities in vitro against M. avium residing within J774, a murine macrophage-like cell line. A conventional passive-entrapment method yielded an encapsulation efficiency of 9% for ciprofloxacin and because of aggregation mediated by the cationic drug, was useful only with liposomes containing < or = 50 mol% negatively charged phospholipid. In contrast, ciprofloxacin was encapsulated with > 90% efficiency, regardless of the content of negatively charged lipids, by a remote-loading technique that utilized both pH and potential gradients to drive drug into preformed liposomes. Both the cellular accumulation and the antimycobacterial activity of ciprofloxacin increased in proportion to the liposome negative charge; the maximal enhancement of potency was 43-fold in liposomes of distearoylphosphatidylglycerol-cholesterol (DSPG-Chol) (10:5). Azithromycin liposomes were prepared as a freeze-dried preparation to avoid chemical instability during storage, and drug could be incorporated at 33 mol% (with respect to phospholipid). Azithromycin also showed enhanced antimycobacterial effect in liposomes, and the potency increased in parallel to the moles percent of negatively charged lipids; azithromycin in DSPG-Chol (10:5) liposomes inhibited intracellular M. avium growth 41-fold more effectively than did free azithromycin. Thus, ciprofloxacin or azithromycin encapsulated in stable liposomes having substantial negative surface charge is superior to nonencapsulated drug in inhibition of M.avium growth within cultured macrophages and may provide more effective therapy of M.avium infections.
Biochemical basis for increased susceptibility to Cidofovir of herpes simplex viruses with altered or deficient thymidine kinase activity
Authors: Mendel D B , Barkhimer D B , Chen M S
Published: June 2012

It has been observed that herpes simplex virus mutants with deficient or altered thymidine kinase activity are more susceptible to Cidofovir (CDV; 1-[(S)-3-hydroxy-2-(phosphonomethoxy)propyl]cytosine dihydrate) in tissue culture than are the parental strains. During infection of cells, the elevation of the dCTP pool by thymidine kinase mutant viruses is less than that induced by the wild-type virus. The competition between CDV diphosphate and dCTP at the viral polymerase is therefore changed in favor of CDV diphosphate, enhancing its activity.
Effects of traditional Chinese medicines on pharmacokinetics of levofloxacin
Authors: Hasegawa T , Yamaki K , Muraoka I , Nadai M , Takagi K , Nabeshima T
Published: June 2012

The effects of single coadministrations of one of three traditional Chinese medicines, Hotyu-ekki-to, Rikkunshi-to, and Juzen-taiho-to, on the pharmacokinetics of levofloxacin (LVFX) were investigated with eight healthy volunteers in an open, random crossover fashion. Subjects each received a single oral dose of LVFX (200 mg) alone and then with a single coadministration of each Chinese medicine. There were no significant differences in any pharmacokinetic parameters of LVFX between the groups. Also, no significant changes in the urinary recovery (> 80%) and renal clearance of LVFX were observed. These results indicate that the Chinese medicines tested have no significant effect on the rate and extent of bioavailability or renal excretion of LVFX.
Macrolide antibiotics inhibit 50S ribosomal subunit assembly in Bacillus subtilis and Staphylococcus aureus
Authors: Champney W S , Burdine R
Published: June 2012

Macrolide antibiotics are clinically important antibiotics which are effective inhibitors of protein biosynthesis in bacterial cells. We have recently shown that some of these compounds also inhibit 50S ribosomal subunit formation in Escherichia coli. Now we show that certain macrolides have the same effect in two gram-positive organisms, Bacillus subtilis and Staphylococcus aureus. Assembly in B. subtilis was prevented by erythromycin, clarithromycin, and azithromycin but not by oleandomycin. 50S subunit formation in S. aureus was prevented by each of seven structurally related 14-membered macrolides but not by lincomycin or two streptogramin antibiotics. Erythromycin treatment did not stimulate the breakdown of performed 50S subunits in either organism. The formation of the 30S ribosomal subunit was also unaffected by these compounds. Assembly was also inhibited in a B. subtilis strain carrying a plasmid with the ermC gene that confers macrolide resistance by rRNA methylation. These results suggest that ribosomes contain an additional site for the inhibitory functions of macrolide antibiotics.
Influence of dexamethasone on efficacy of ceftriaxone and vancomycin therapy in experimental pneumococcal meningitis
Authors: Cabellos C , Martinez-Lacasa J , Martos A , Tubau F , Fernández A , Viladrich P F , Gudiol F
Published: June 2012

Using a rabbit model of meningitis, we sought to determine whether concomitant use of dexamethasone affects the penetration and efficacy of ceftriaxone or vancomycin in cerebrospinal fluid. Rabbits were inoculated with a penicillin-sensitive strain of Streptococcus pneumoniae and treated with ceftriaxone or vancomycin with or without dexamethasone. In the ceftriaxone-treated groups, no statistically significant differences were seen between the group treated with dexamethasone and that without dexamethasone; however, in the vancomycin-treated groups we found statistically significant lower cerebrospinal fluid vancomycin levels at 2 h in the dexamethasone-treated rabbits and differences in bacterial killing.
Sequence and analysis of the rpoB gene of Mycobacterium smegmatis
Authors: Hetherington S V , Watson A S , Patrick C C
Published: June 2012

The rpoB gene encodes the beta subunit of the DNA-dependent RNA polymerase of bacteria. Mutations in defined areas result in resistance to rifampin. Mycobacterium smegmatis is naturally resistant to rifampin, but analysis of the rpoB gene revealed no identifiable rifampin resistance mutations. Another mechanism of resistance may be present.
Antigen-Specific Memory T Cell Responses after Vaccination with an Oral Killed Cholera Vaccine in Bangladeshi Children and Comparison to Responses in Patients with Naturally Acquired Cholera
Authors: Arifuzzaman Mohammad , Rashu Rasheduzzaman , Leung Daniel T. , Hosen M. Ismail , Bhuiyan Taufiqur Rahman , Bhuiyan M. Saruar , Rahman Mohammad Arif , Khanam Farhana , Saha Amit , Charles Richelle C. , LaRocque Regina C. , Weil Ana A. , Clements John D. , Holmes Randall K. , Calderwood Stephen B. , Harris Jason B. , Ryan Edward T. , Qadri Firdausi
Published: June 2012

Young children, older children, and adults develop comparable levels and durations of immunity following cholera. In comparison, young children receiving oral killed cholera vaccines (OCV) develop a lower level and shorter duration of protection than those of older children and adults. The reasons for this are unclear. We investigated OCV-induced memory T cell responses in younger and older children and compared responses to those in children with cholera. We found that patients with cholera developed significant levels of toxin-specific effector memory T cells (TEM) with follicular helper and gut-homing characteristics. Older children (6 to 14 years of age) receiving two doses of OCV containing recombinant cholera toxin B subunit (rCTB) had more modest TEMresponses with follicular helper and gut-homing characteristics, but younger vaccinees (24 to 71 months of age) did not develop TEMresponses. The TEMresponse correlated positively with subsequent IgG memory B cell responses specific to rCTB in older vaccinees. Cytokine analyses indicated that cholera patients developed significant Th1, Th17, and Th2 responses, while older children receiving vaccine developed more modest increases in Th1 and Th17 cells. Younger vaccinees had no increase in Th1 cells, a decrease in Th17 cells, and an increase in regulatory T (Treg) cells. Our findings suggest that T cell memory responses are markedly diminished in children receiving OCV, especially young children, compared to responses following naturally acquired cholera, and that these differences affect subsequent development of memory B cell responses. These findings may explain the lower efficacy and shorter duration of protection afforded by OCV in young children.
Nucleoside Analogue Mutagenesis of a Single-Stranded DNA Virus: Evolution and Resistance
Authors: Domingo-Calap Pilar , Pereira-Gómez Marianoel , Sanjuán Rafael
Published: June 2012

It has been well established that chemical mutagenesis has adverse fitness effects in RNA viruses, often leading to population extinction. This is mainly a consequence of the high RNA virus spontaneous mutation rates, which situate them close to the extinction threshold. Single-stranded DNA viruses are the fastest-mutating DNA-based systems, with per-nucleotide mutation rates close to those of some RNA viruses, but chemical mutagenesis has been much less studied in this type of viruses. Here, we serially passaged bacteriophage ϕX174 in the presence of the nucleoside analogue 5-fluorouracil (5-FU). We found that 5-FU was unable to trigger population extinction for the range of concentrations tested, but it negatively affected viral adaptability. The phage evolved partial drug resistance, and parallel nucleotide substitutions appearing in independently evolved lines were identified as candidate resistance mutations. Using site-directed mutagenesis, two single-nucleotide substitutions in the lysis protein E (T572C and A781G) were shown to be selectively advantageous in the presence of 5-FU. In RNA viruses, base analogue resistance is often mediated by changes in the viral polymerase, but this mechanism is not possible for ϕX174 and other single-stranded DNA viruses because they do not encode their own polymerase. In addition to increasing mutation rates, 5-FU produces a wide variety of cytotoxic effects at the levels of replication, transcription, and translation. We found that substitutions T572C and A781G lost their ability to confer 5-FU resistance after cells were supplemented with deoxythymidine, suggesting that their mechanism of action is at the DNA level. We hypothesize that regulation of lysis time may allow the virus to optimize progeny size in cells showing defects in DNA synthesis.
Pathogenic Consequences of Vaginal Infection with CCR5-Tropic Simian-Human Immunodeficiency Virus SHIV
SF162P3N
Authors: Shakirzyanova Madina , Tsai Lily , Ren Wuze , Gettie Agegneu , Blanchard James , Cheng-Mayer Cecilia
Published: June 2012

We previously reported efficient transmission of the pathogenic R5 simian-human immunodeficiency virus SHIV
SF162P3N
isolate in Indian rhesus macaques by intravenous and intrarectal inoculations, with a switch to CXCR4 coreceptor usage in ∼50% of infected animals that progressed rapidly to disease. Since women continue to be disproportionately affected by HIV, we developed an animal model based on the intravaginal challenge of female rhesus monkeys with SHIV
SF162P3N
and sought to validate the utility of this model to study relevant aspects of HIV transmission and pathogenesis. The effect of viral dose on infection outcome was evaluated to determine the optimal conditions for the evaluation of HIV-1 preventive and therapeutic strategies. We found that the virus can successfully cross the vaginal mucosal surface to establish infection and induce disease with coreceptor switch, but with lower efficiencies compared to intravenous and rectal transmissions. In contrast to intrarectal infection, peak and cumulative viral load over a 1 year-infection period were significantly greater in macaques exposed intravaginally to lower rather than higher inoculum doses. Moreover, low and transient viremia was observed only in macaques that were challenged intravaginally twice within the same day with a high dose of virus, which can be seen as doubling the dose. Taken together, these results show that SHIV
SF162P3N
can successfully transmit across the genital mucosa, undergo coreceptor switch, and induce disease. However, the administered dose appears to impact SHIV
SF162P3N
vaginal infection outcome in an unexpected manner.
Quantitative Measurement of Human Papillomavirus Type 16 E5 Oncoprotein Levels in Epithelial Cell Lines by Mass Spectrometry
Authors: Sahab Ziad , Sudarshan Sawali R. , Liu Xuefeng , Zhang YiYu , Kirilyuk Alexander , Kamonjoh Christopher M. , Simic Vera , Dai Yuhai , Byers Stephen W. , Doorbar John , Suprynowicz Frank A. , Schlegel Richard
Published: June 2012

The high-risk human papillomavirus type 16 (HPV-16) E5 protein (16E5) induces tumors in a transgenic mouse model and may contribute to early stages of cervical carcinogenesis. Although high-risk E5 expression is generally thought to be lost during the progression to cervical carcinoma following integration of HPV DNA into the host genome, episomal viral DNA has been documented in a subset of HPV-16-positive malignant lesions. Numerous studies have shown that transcripts that could potentially encode 16E5 are present in cervical biopsy specimens and cervical cancer cell lines, but the presence of E5 protein has been demonstrated in only two reports that have not been corroborated. In the present study, we show that trypsin cleavage of 16E5 generates a unique four-amino-acid C-terminal peptide (FLIT) that serves as a marker for E5 expression in transfected cells and epithelial cell lines containing integrated and episomal HPV-16 DNA. Following trypsin cleavage, reversed-phase chromatography and mass spectrometry (MS) were used to detect FLIT. Immunoprecipitation assays using a newly generated anti-16E5 antibody confirmed that 16E5 was solely responsible for the FLIT signal, and deuterated FLIT peptide provided an internal standard that enabled us to quantify the number of 16E5 molecules per cell. We show that 16E5 is expressed in the Caski but not in the SiHa cervical cancer cell line, suggesting that 16E5 may contribute to the malignant phenotype of some cervical cancers, even in cells exclusively containing an integrated HPV genome.
Restricted Varicella-Zoster Virus Transcription in Human Trigeminal Ganglia Obtained Soon after Death
Authors: Ouwendijk Werner J. D. , Choe Alexander , Nagel Maria A. , Gilden Don , Osterhaus Albert D. M. E. , Cohrs Randall J. , Verjans Georges M. G. M.
Published: June 2012

We analyzed the varicella-zoster virus (VZV) transcriptome in 43 latently infected human trigeminal ganglia (TG) with postmortem intervals (PMIs) ranging from 3.7 to 24 h. Multiplex reverse transcriptase PCR (RT-PCR) revealed no VZV transcripts with a PMI of <9 h. Real-time PCR indicated a significant increase (
P
= 0.02) in VZV ORF63 transcript levels but not the virus DNA burden with longer PMI. Overall, both the breadth of the VZV transcriptome and the VZV ORF63 transcript levels in human cadaver TG increased with longer PMI.
Reverse Genetics Analysis of Poxvirus Intermediate Transcription Factors
Authors: Warren Robin D. , Cotter Catherine A. , Moss Bernard
Published: June 2012

Vaccinia virus transcription is regulated in three stages. An intermediate transcription factor, comprised of virus-encoded polypeptides A8 and A23, was previously identified by
in vitro
analyses. To investigate its role, we engineered cells that stably expressed both subunits and complemented the replication of A8 and A23 deletion mutant viruses. Without A8 or A23, viral early gene expression and DNA replication occurred but intermediate and late gene expression and resolution of genome concatemers were not detected.
A modern approach to equine dentistry 4. Routine treatments
Authors: Tremaine Henry , Pearce Chris
Published: June 2012

This article, the final one in a series of four to review recent developments in equine dentistry, describes the equipment and routine techniques required for the maintenance of oral health in horses, with the aim of preserving normal masticatory function, and provides an approach to the treatment of dental lesions and periodontal disease. Part 1, published in the January issue of In Practice (volume 34, pages 2–10), described the procedure for a routine dental examination in horses, while Part 2, published in the February issue (volume 34, pages 78–89), highlighted the common dental and oral lesions that may be seen in this species. Part 3, published in the March issue (volume 34, pages 114–127), described the use of imaging to aid the diagnosis of such lesions.
Fading puppies – reality or myth?
Authors: Blunden Tony
Published: June 2012

The term fading puppy syndrome (FPS) or fading puppy complex has been used inconsistently to refer to any failure‐to‐thrive syndrome beginning from birth to weaning that results in death despite all attempts at treatment. However, the term ‘fading’ has been mostly applied to neonatal puppies. The neonatal period has been defined variously as the first seven or 10 or 21 days. The reality is that the neonate is born immature and maturation occurs only gradually over a period of weeks. For example, the external ear canals of the puppy are occluded until between 12 and 14 days after birth. Further enlargement of the canal continues until its size and relationship to the other external ear structures appears to be normal, at about five weeks of age. Glomerular filtration in the kidneys increases from 21 per cent at birth to 53 per cent at eight weeks old. Tubular secretion also matures at eight weeks of age. Thus, any cut‐off point in development in the first weeks of life is arbitrary but the change from the nest period, when the neonate is completely reliant on the dam for its survival, to the stage when it can start to feed itself and walk away from the nest marks a turning point when the puppy becomes far less vulnerable to a variety of factors.
The use of pollination networks in conservation1This article is part of a Special Issue entitled “Pollination biology research in Canada: Perspectives on a mutualism at different scales”.
Authors: Elle Elizabeth , Elwell Sherri L. , Gielens Grahame A.
Published: June 2012

Recent concern about declines in pollinating insects highlights the need for better understanding of plant–pollinator interactions. One promising approach at the community scale is network analysis, which allows actual interactions to be assessed, unlike biodiversity surveys, which only identify the potentially interacting organisms. We highlight useful network properties for conservation research and examples of their use in the study of rare species, invasive species, responses of communities to climate change, and habitat loss and restoration. We suggest that nestedness, degree, and interaction strength asymmetry are the most useful network properties for applied research on plant–pollinator interactions, but also highlight practical concerns regarding their measurement. We encourage the adoption of a network approach when an understanding of function within communities, rather than simple community composition, is useful for management.
Biostratigraphic, strontium isotopic, and geologic constraints on the landward movement and fragmentation of terranes within the Tofino Basin, British Columbia
Authors: Johns Marjorie J. , Trotter Julie A. , Barnes Christopher R. , Narayan Y. Roshni
Published: June 2012

Significant advancements in understanding the complex evolution of the Tofino Basin at a convergent accretionary margin are enabled by combining contextual geologic information with new isotopic and paleontological data. A high-resolution Cenozoic chronostratigraphy of the basin is constrained by strontium isotope ages (36.9–1.3 Ma) of Late Eocene to Pleistocene foraminifers together with a revised biostratigraphy (foraminifers and ichthyoliths) from six offshore wells and outcrop samples, new specimen thermal alteration values, and existing well log data. These data are integrated with archival multichannel seismic and magnetic data to interpret offshore well positions with relation to sub-basins and structural highs of the Pacific Rim and Crescent terranes, and other accreted strata. Six regions of the Tofino Basin are defined based on structure and depositional differences during the Eocene to Holocene history of accretion and fragmentation of the Crescent terrane and it underthrusting the Pacific Rim terrane. Subsequent oceanic sediment accretions and deposition of overlying sediments up to about 4000 m thick resulted as the Juan de Fuca plate subducted beneath Vancouver Island. Observations include different fragmentations and landward movements of the Crescent and Pacific Rim terranes in the regions and two fault styles in the Ucluelet and Carmanah regions where six new sub-basins are defined. Results, especially for the Ucluelet and Carmanah sub-basins, indicate periods of deformation during the Late Eocene, Late Oligocene, Middle–Late Miocene, and post middle Pliocene, whereas the Early Oligocene and Early Miocene had periods of relatively slow and less disturbed deposition.
Seismic force demand on ductile reinforced concrete shear walls subjected to western North American ground motions: Part 1 — parametric study
Authors: Boivin Yannick , Paultre Patrick
Published: June 2012

A parametric study of regular ductile reinforced concrete (RC) cantilever walls designed with the 2010 National building code of Canada and the 2004 Canadian Standards Association (CSA) standard A23.3 for Vancouver is performed to investigate the influence of the following parameters on the higher mode amplification effects, and hence on the seismic force demand: number of storeys, fundamental lateral period (T), site class, wall aspect ratio, wall cross-section, and wall base flexural overstrength (γw). The study is based on inelastic time-history analyses performed with a multilayer beam model and a smeared membrane model accounting for inelastic shear–flexure–axial interaction. The main conclusions are that (i) T and γware the studied parameters affecting the most dynamic shear amplification and seismic force demand, (ii) the 2004 CSA standard A23.3 capacity design methods are inadequate, and (iii) a single plastic hinge design may be inadequate and unsafe for regular ductile RC walls with γw < 2.0.
Performance-based mix design of unmodified and lime-modified hot mix asphalt
Authors: Lee Sangyum , Baek Cheolmin , Park Je-Jin
Published: June 2012

This paper presents the performance evaluation of unmodified and lime-modified hot mix asphalt (HMA) mixtures at varying asphalt content using asphalt mixture performance test developed from National Cooperative Highway Research Program project 9-19 and 9-29 and the viscoelastic continuum damage finite element analysis. Test methods adopted in this study are the dynamic modulus test for stiffness, the triaxial repeated load permanent deformation test for rutting, and the direct tension test for fatigue cracking. The findings from this study support conventional understanding of the effects of asphalt content and lime modification on the fatigue cracking and rutting performance. Finally, the optimum asphalt content for both lime-modified and unmodified mixtures are proposed based on the knowledge gleaned from the performance-based mix design methodology. With additional validation and calibration, the comprehensive methodology described in this paper may serve as the foundation for a performance-based HMA mix design and performance-related HMA specifications.
Bond characteristics of steel fibre reinforced self-compacting concrete
Authors: Aslani Farhad , Nejadi Shami
Published: June 2012

Steel fibre reinforced self-compacting concrete (SFRSCC) is a relatively new composite material that combines the benefits of the self-compacting concrete (SCC) technology with the advantages derived from the fibre addition to a brittle cementitious matrix. Steel fibres improve many of the properties of SCC elements including tensile strength, ductility, toughness, energy absorption capacity, fracture toughness and cracking. Although the available research regarding the influence of steel fibres on the properties of SFRSCC is limited, this paper investigates the bond characteristics between steel fibre and SCC. Based on the available experimental results, the current analytical steel fibre pullout model is modified by considering the different SCC properties and different fibre types (smooth, hooked) and fibre inclination. To take into account the effect of fibre inclination in the pullout model, apparent shear strengths (τ(app)) and slip coefficient (β) are incorporated to express the variation of pullout peak load and the augmentation of peak slip as the inclined angle increases. These variables are expressed as functions of the inclined angle (ϕ).