Co-occurrence of mcr-1 in the chromosome and on an IncHI2 plasmid: persistence of colistin resistance in Escherichia coli

Two colistin-resistant Escherichia coli strains (FS13Z2S and FS3Z6C) possessing chromosomally encoded mcr-1 isolated from swine were characterised. Whole-genome sequencing revealed that in strain FS13Z2S mcr-1 occurred in triplicate in the chromosome with another copy encoded on a pHNSHP45-2-like IncHI2 plasmid, whereas in strain FS3Z6C only one copy mcr-1 was inserted in the chromosome. It seems likely that the triplication of chromosomal copies of mcr-1 in FS13Z2S is due to intramolecular transposition events via a composite transposon containing an mcr-1 cassette bracketed by two copies of insertion sequence ISApl1, and the pap2 gene at the insertion site was truncated by an IS1294-like element. In plasmid pFS13Z2S and the chromosome of strain FS3Z6C, only a single copy of ISApl1 was present upstream of the mcr-1 cassette. The two strains exhibited similar colistin minimum inhibitory concentrations (MICs) and featured phosphoethanolamine addition to lipid A, without regard to the copy number of mcr-1. The mcr-1-harbouring plasmid was unstable in wild-type strain FS13Z2S and was quickly lost after 7 days of passage on colistin-free Luria–Bertani broth containing 0.5% SDS, but the mcr-1 copies on the chromosome persisted. These results reveal that the single copy of mcr-1 could result in modification of lipopolysaccharide (LPS) and cause colistin resistance in E. coli. Acquisition of multiple copies of mcr-1, especially on the chromosome, would facilitate stable persistence of colistin resistance in the host strain.     

Release of large amounts of lipopolysaccharides from Pseudomonas aeruginosa cells reduces their susceptibility to colistin

Pseudomonas aeruginosa is an important etiological agent of opportunistic infections. Injectable colistin is available as a last-line treatment option for multidrug-resistant P. aeruginosa infections. When cells were inoculated at a high number, colistin-susceptible P. aeruginosa grew on agar medium containing colistin at a concentration 10-fold higher than the minimum inhibitory concentration without acquiring colistin resistance. This study examined the responsible mechanism for growth in the presence of a high concentration of colistin. Cell wash fluid derived from P. aeruginosa efficiently reduced colistin antimicrobial activity. This reduction was mediated by lipopolysaccharide (LPS) in the wash fluid. Extracellular LPS inhibited colistin activity more effectively than cell-bound LPS in fixed cells. Cell wash fluids from Escherichia coli and Acinetobacter baumannii also reduced colistin activity; however, they were less potent than those from P. aeruginosa. The amount of LPS in cell wash fluid from P. aeruginosa was approximately 10-fold higher than that in fluid from E. coli or A. baumannii. In conclusion, cell-free LPS derived from bacterial cells inhibited the antimicrobial activity of colistin, and this effect was greatest for P. aeruginosa. Thus, large amounts of broken and dead cells of P. aeruginosa at infection foci will reduce the effectiveness of colistin, even against cells that have not yet acquired resistance.     

Comparative effects of Ca-ethylenediaminetetraacetic acid (EDTA), ZnEDTA,and ZnCaEDTA in mobilizing lead

Male Long-Evans rats weighing approximately 240 g were given (Pb) at 14 mg/kg as the acetate by slow iv infusion 17 days prior to chelate treatment. The chelating agents were administered by continuous iv infusion at either 1 mmol/kg over 6 hr or 6 mmol/kg over 24 hr or at 0.16 mmol/kg/day by sc injections. ZnEDTA was 60% and ZnCaEDTA 76% as effective as CaEDTA in promoting urinary Pb excretion at 1 mmol/kg over 6 hr, iv. At 6 mmol/kg/24 hr, iv, ZnEDTA was 76% and ZnCaEDTA 98% as effective as CaEDTA. Mean urinary Pb excretion for each chelate via the sc route and the lowest iv route of administration was the same. Blood δ-aminolaevulinic acid dehydratase (ALAD) activity (an indicator of Pb toxicity) was enhanced approximately twofold by CaEDTA, two and one-half-fold by ZnCaEDTA, and fivefold by ZnEDTA treatment. It is suggested that the safety of EDTA can be markedly enhanced if administered as a ZnCaEDTA chelate without appreciably diminishing its efficacy in promoting urinary Pb excretion.     

Effects of food dyes on Paramecium caudatum: toxicity and inhibitory effects on leucine aminopeptidase and acid phosphatase activity.

The toxic effect of 14 food dyes was studied in Paramecium caudatum. It was found that xanthene dyes containing halogen atoms in their molecules were more toxic than other groups of food dyes. Phloxin and rose bengal containing chlorine were especially toxic. The effect of food dyes on leucine aminopeptidase, acid phosphatase, and γ-glutamyl transpeptidase activity in P. caudatum was studied in order to investigate the mechanism of toxicity. Phloxin and rose bengal inhibited leucine aminopeptidase remarkably. The inhibitory effect of food dyes on leucine aminopeptidase in vitro is consistent with the toxic effect of the dyes on the survival time of P. caudatum. A possible correlation between toxicity and inhibition of the activity of enzymes involved in the digestive process is discussed.     

Wide dissemination of SCCfusC in fusidic acid-resistant coagulase-negative staphylococci and implication for its spread to methicillin-resistant staphylococcus aureus in Taiwan

The fusidic acid (FUS) resistance determinants fusB, fusC, fusD and fusF in coagulase-negative staphylococci (CoNS) clinical isolates were examined. Among 208 FUS-resistant isolates, the fusB gene was the most common resistance determinant in each species, except in Staphylococcus hominis subsp. hominis or in species carrying intrinsic fusD or fusF. In S. hominis subsp. hominis, the fusC gene was the major determinant responsible for FUS resistance. To understand the genetic context of fusC in S. hominis subsp. hominis, 31 fusC-positive S. hominis subsp. hominis isolates were examined. Among these isolates, 14 carried SCCfusC, 3 carried an SCC₄₇₆-like element and 7 carried a new SCC structure (SCC₃₃₉₀). As shown by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) analyses, the S. hominis subsp. hominis clinical isolates showed limited clonality. Taken together, SCCfusC has been found in S. hominis subsp. hominis, Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus capitis subsp. ureolyticus and Staphylococcus aureus, suggesting its wide distribution and spread among different species of staphylococci.     

Measuring the quality and completeness of medication-related information derived from hospital electronic health records database

ObjectiveElectronic Health Records (EHRs) database is a great source for pharmacoepidemiological research as thousands of patients’ clinical and medication information is stored in the database. However, the use of EHRs database for research purposes depends greatly on the accuracy and completeness of the data being used. This study mainly aimed to assess the completeness of EHRs patients’ medication-related information.DesignA retrospective cross-sectional study using data extracted from the EHRs database was conducted.SettingThe EHRs data was obtained from a single tertiary hospital in Saudi Arabia.Main outcome measure(s)The completeness of data was measured considering if a patients’ record contains all desired types of data (i.e., patients’ demographics, clinical diagnosis, and medication-related information).ResultsA total of 23,411 unique individuals were identified after extracting the data from the EHRs. The study found that 89.9% of the patients had a complete data (i.e., age, gender, marital status, nationality, encounter type, and clinical diagnosis). Further, 83.1% of the patients had complete medication-related information. Subgroup analysis by the encounter type indicated that the data was 91.0% complete for outpatient encounter and 93.2% complete for inpatient encounter.ConclusionThe study findings indicate that the completeness of the data varies by the desired types of data. EHRs can be a potentially great resource to conduct research to assess medication use. Further studies focusing on the content and completeness of EHRs for a specific patient population and evaluate other dimensions of EHRs data quality are needed.     

A possible protective role for reduced glutathione in aflatoxin B1 toxicity: effect of pretreatment of rats with phenobarbital and 3-methylcholanthrene on aflatoxin toxicity.

To determine if brain-acetylcholinesterase (AChE) inhibition in a marine teleost Lagodon rhomboides (pinfish) by an organophosphate pesticide (naled) is specific enough to diagnose anticholinesterase poisoning, brain-AChE inhibition by sublethal exposure in seawater was compared to brain-AChE inhibition caused by lethal exposure. A sublethl exposure did not inhibit brain-AChE as much as lethal exposure in periods of 24, 48, and 72 hr. Consistent levels of inhibition (84–89% inhibition) occurred when 40–60% of an exposed population of pinfish was killed. This correlation of brain-AChE inhibition with exposure and death in a fish population shows that brain-AChE measurements are of value in diagnosing anticholinesterase poisoning in a marine fish.     

Physiological role of nitric oxide for regulation of arterial stiffness in anesthetized rabbits

We assessed effects of acetylcholine and Nω-Nitro-l-arginine methyl ester hydrochloride (l-NAME) on the cardio-ankle vascular index (CAVI), an indicator of arterial stiffness from origin of aorta to tibial artery, in halothane-anesthetized rabbits. Acetylcholine decreased the blood pressure, femoral vascular resistance and CAVI, whereas l-NAME did not affect the CAVI at a hypertensive dose. The acetylcholine-induced decrement of CAVI was completely suppressed by l-NAME. These results suggest that the arterial stiffness in rabbits may be independent from homeostatic production of nitric oxide, however, it can be decreased by large amounts of nitric oxide that are intrinsically produced by exogenously administered acetylcholine.     

Combined citicoline and docosahexaenoic acid treatment improves cognitive dysfunction following transient brain ischemia

Phospholipids are structural components of cellular membranes that play important roles as precursors for various signaling pathways in modulating neuronal membrane function and maintenance of the intracellular environment. Phosphatidylcholine (PtdCho) is the most abundant cellular phospholipid. Citicoline and docosahexaenoic acid (DHA) are essential intermediates in the synthesis of PtdCho. Both PtdCho intermediates have independently shown neuroprotective effects in cerebral ischemia, but their combined effect is unknown. This study aimed to investigate the combined effect of oral citicoline and DHA treatment on improvement of cognitive deficits following cerebral ischemia using a 20-min bilateral common carotid artery occlusion (BCCAO) mouse model. BCCAO ischemic mice were treated for a total of 11 days with a combination of citicoline (40 mg/kg body weight/day) and DHA (300 mg/kg body weight/day) or each alone. Combined citicoline and DHA synergistically and significantly improved learning and memory ability of ischemic mice compared with either alone. Further, citicoline and DHA treatment significantly prevented neuronal cell death, and slightly increased DHA-containing PtdCho in the hippocampus, albeit not significantly. Taken together, these findings suggest that combined citicoline and DHA treatment may have synergistic benefits for partially improving memory deficits following transient brain ischemia.     

Electropharmacological effects of intracellular Ca2+ handling modulator caldaret on the heart assessed in the halothane-anesthetized dogs

We analyzed how the enhancement of net sarcoplasmic reticulum (SR) Ca²⁺ uptake may affect cardiac electrophysiological properties in vivo by using caldaret which can decrease SR diastolic Ca²⁺ leak, enhance SR Ca²⁺ reuptake and inhibit reverse-mode Na⁺/Ca²⁺ exchanger. Caldaret in doses of 0.5, 5 and 50 μg/kg was intravenously administered over 10 min to the halothane-anesthetized beagle dogs (n = 5), attaining pharmacologically active plasma concentration. The low and middle doses of caldaret increased the ventricular contraction, which could be explained by its on-target pharmacological activities. The high dose enhanced the sinus automaticity followed by its suppression in addition to the increase of the total peripheral resistance, which may be unfavorable for treating diastolic heart failure. The low and middle doses enhanced the atrioventricular conduction, which may have some potential for predisposing the atria to the onset of atrial fibrillation via an induction of mitral and/or tricuspid regurgitation. The middle and high doses of caldaret prolonged the ventricular effective refractory period without altering the intraventricular conduction or repolarization period, which may prevent the onset of ventricular arrhythmias. Thus, modulation of intracellular Ca²⁺ handling by caldaret can induce not only inotropic effect, but also various electrophysiological actions on the in situ heart.     

Effects of age and sex on retention of mercury by methyl mercury-treated rats

Male and female Long Evans rats 7, 15, 20, 24, or 56 days old received a single subcutaneous injection of 1 μmol of methyl mercury-203/kg and the whole body retention of radiomercury was determined for up to 139 days thereafter. For rats dosed at 7 or 15 days of age, whole body clearance of mercury was extremely slow until animals reached 17 to 18 days of age. Subsequent excretion was monoexponential in the 7-day-old group and biexponential in the 15-day-old group. For rats dosed at 20, 24, or 56 days of age, onset of excretion was immediate and the pattern of clearance was biexponential. In rats dosed at 56 days of age, the retention of mercury by the average male and average female was significantly different (p = 0.001). No sexual difference in the estimate of whole body retention of mercury was seen in the other age groups. The presence of an interval of very slow excretion of mercury in young rats and the subsequent slower excretion of mercury in these animals than in rats dosed with methyl mercury later in life suggest that increased hazards of methyl mercury exposure in early life may be related to increased retention of the organomercurial or inorganic Hg.     

Effect of pentachloronitrobenzene upon egg production, hatchability, and residue accumulation in the tissues of White Leghorn hens.

Individual groups of White Leghorn female chickens were fed graded concentrations (0, 10, 50, 100, and 1000 ppm) of pentachloronitrobenzene (PCNB), containing hexachlorobenzene (HCB), pentachlorobenzene (PCB), and tetrachloronitrobenzene (TCNB) as contaminants, from 1 day through 35 weeks of age. No concentration of PCNB caused toxic effects or death. Histopathologic examination of 10 organs, including ovaries, failed to reveal abnormalities in either control or treated groups. Onset of egg production was delayed for 1 month, and hatchability of eggs was significantly lower in hens at 1000 ppm PCNB. No embryonic abnormalities were observed. Only small amounts of PCNB and metabolites, pentachloroaniline (PCA) and pentachloro-phenylmethylsulfide (PCMS), were found in eggs. Conversely, HCB and PCB were readily transmitted into eggs. TCNB either was not detected or was present in only trace or finite amounts in eggs. PCNB, contaminates of PCNB (HCB, PCB, and TCNB), and metabolites of PCNB (PCA and PCMS) occurred in the highest concentration in adipose tissue. PCNB was not detectable (<0.05 ppm) in adipose tissue by 6 days following its withdrawal from the diet. In egg, the half-life of PCNB was not determined since only trace amounts were detected. Depletion half-lives of HCB and PCB in eggs were as high as 5.1 and 3.6 weeks, respectively.     

Testosterone-stimulation of adenyl cyclase and cyclic 3',5'-adnosine monophosphate- 3 H formation in rat seminal vesicles

Seminal vesicles obtained from normal or castrate rats were capable of converting radioactive adenosine into labeled cyclic 3′,5′-adenosine monophosphate (cAMP-³H). Addition of testosterone (10^?6M) in vitro caused about a 60 per cent stimulation in the levels of vesicular cAMP-³H. Castration (96 hr) led to a significant decrease in the content of cAMP-³H and adenyl cyclase activity of this accessory organ; it did not significantly decrease the concentration. A single injection of testosterone to castrate rats produced a marked enhancement of vesicular adenyl cyclase (96 hr later). These results support the concept that stimulation of the cAMP-adenyl cyclase system is associated with the mechanism of action of androgens on their target cells.     

Effect of α-fluoromethylhistidine,a suicide inhibitor of histidine decarboxylase,on histamine levels in mouse tissues

The effects of α-fluoromethylhistidine (α-FMH), a new suicide inhibitor [Kollonitsch et al., Nature, Lond.274, 906 (1978)], on histidine decarboxylase (HDC) activities and histamine contents of the skin, fundic stomach and brain of mice were investigated. Four hours after i.p. administration of α-FMH to ddy mice, HDC activities in the brain, stomach and skin had decreased in a dose-dependent way (1–25 mg/kg), by a maximum of 90–95%. The histamine levels in the brain and stomach decreased to 50% of the control levels, whereas the level in the skin did not change at all. The time courses of changes in HDC activities and histamine levels were examined. After i.p. administration of 25 mg/kg of α-FMH, HDC activities in these tissues dropped rapidly within 1 hr. Recovery of HDC activities in the stomach and skin began within 12 hr, but the activity in the brain remained low for 24 hr, confirming the result of Garbarg et al. [J. Neurochem.35, 1045 (1980)]. The histamine content of the stomach decreased to 40% of the original level in 8 hr and recovered within 12 hr, whereas that in the brain decreased to 50% and remained low for more than 24 hr. The histamine content of the skin did not change. These results suggest that the histamine level that was not reduced by α-FMH was derived from mast cells. During the above experiments, no behavioral changes of the animals were detected. α-FMH prevented the increase in HDC activity in mouse kidney on day 18 of gestation when administered i.p. every 12 hr from day 13. No abnormalities were seen in fetuses and neonates after this treatment. It is concluded that α-FMH causes depletion of newly synthesized histaminein situ and, thus, is useful for studies on histamine.