Promoting veterinary medication safety - Exploring the competencies of community pharmacy professionals in veterinary pharmacotherapy.

The science of veterinary medicine is currently lacking studies on medication safety, although its importance in protecting animals from medication errors is central. Pharmacy professionals have an important role in ensuring medication safety of both prescription and over-the-counter medications of animals. However, this requires adequate competencies of pharmacy professionals in veterinary pharmacotherapy. The present study aimed to explore the competencies of pharmaceutical staff in community pharmacies in veterinary pharmacotherapy, which factors influence these competencies and what kind of information sources they typically use on veterinary pharmacotherapy. The study was conducted as a cross-sectional online survey targeted to pharmacy professionals in the Finnish community pharmacies, providing 596 responses. Less than half of the respondents (41%, n = 246) are considered to possess good competencies in veterinary pharmacotherapy. A third of the respondents (35%, n = 211) would dispense an anti-inflammatory drug for an animal off-label, whereas 24% (n = 145) would not interview the pet owner to discover the need for internal parasite medication before dispensing the drug. A small proportion (<1%, n = 5) would have dispensed a broad-spectrum internal parasite medication. Approximately a quarter of the respondents (27%, n = 159) stated that they acquired information on pharmacotherapy only from the material produced by the manufacturers of veterinary drugs. The competencies of pharmacy professionals in veterinary pharmacotherapy need to be strengthened in many areas to better promote veterinary medication safety. It should also be ensured that pharmacy professionals can access and use independent, high-quality information on veterinary pharmacotherapy.

Student selection in radiography education. A narrative review.

OBJECTIVES: Due to effects on study success, radiography student selection has a major impact on higher education institutions and applicants. However, there is very little research to demonstrate which selection methods and contents are most successful in radiography education. This study aimed to describe the methods and contents used in radiography student selection and factors related to study success. KEY FINDINGS: A narrative review was undertaken. A computerized search in four databases limited to studies published between January 2000 and June 2021. Ten quantitative, mainly retrospective, studies were included. The review identified 23 selection methods; of these, interview (n = 4), Scholastic Aptitude Test (n = 3), American College Test (n = 2) and reference letter (n = 2) were used more than once in radiography student selection. The content of the selection methods was identified in four categories including 44 factors. The most often assessed content was category of learning skills while the least often assessed concerned categories of social skills, personality traits and career choice. Regarding study success, factors of learning skills, namely mathematics, physics, biology, anatomy, physiology, natural sciences, a composite of factors comprising electronics and a composite of factors comprising mechanics predicted study success. Factors of social skills, personality traits and career choice were not related to study success. CONCLUSION: The methods used and contents assessed vary greatly in radiography student selection. The results suggest using the content in the four categories in the selection of radiography students. IMPLICATIONS FOR PRACTICE: Further research is needed to clarify the methods, with knowledge of the reliability and validity and the contents for the suggested categories, and to demonstrate their relationship to study success and identify the core content of radiography student selection especially in European context.

Desflurane induces more cerebral vasodilation than isoflurane at the same A-line autoregressive index level.

BACKGROUND: Clinical use of desflurane in neuroanesthesia remains under debate. Comparison of dose-dependent vasodilatory properties between desflurane and isoflurane, the more traditional volatile agent for clinical neuroanesthesia, requires equianesthetic dosing of the agents. Reproducible neurophysiological measurements of the level of anesthesia in an individual, e.g. the A-line autoregressive index (AAI), can be used for an equipotent dosage of two volatile agents in the same individual. METHODS: Desflurane and isoflurane, in randomized order, were titrated to a stable AAI level of 15-20 in 18 ASA I or II patients. The mean flow velocity (Vmca) and pulsatility index (PI) in the middle cerebral artery were then measured with transcranial Doppler at an end-tidal CO(2) concentration of 4.4%. RESULTS: For desflurane Vmca was 11% higher [95% confidence interval (CI), 5-18%; P = 0.0020] and PI was 13% lower (95% CI, 3-23%; P = 0.0083) than for isoflurane. The mean arterial blood pressure did not differ between the agents. The fraction of MAC necessary for the intended AAI level was 35% lower (95% CI, 20-49%; P = 0.00016) with desflurane than with isoflurane. CONCLUSION: Desflurane was associated with more cerebral vasodilation than isoflurane at the same depth of anesthesia, as indicated by the AAI. This attributes further reason for caution in the use of desflurane in clinical neuroanesthesia. The difference between desflurane and isoflurane in the MAC fractions required for the same AAI level confirms the limitations of MAC in defining the level of anesthesia.

Sevoflurane induces less cerebral vasodilation than isoflurane at the same A-line autoregressive index level.

BACKGROUND: The use of sevoflurane in neuroanesthesia is still under debate. Comparison of dose-dependent vasodilatory properties between sevoflurane and isoflurane, the more traditional neuroanesthetic agent, requires comparable dosing of the agents. A-line autoregressive index (AAI) provides reproducible individual measurement of anesthetic depth. METHODS: Sevoflurane and isoflurane, in randomized order, were titrated to a stable AAI of 15-20 in each of 18 ASA I or II patients. The mean flow velocity (Vmca) and pulsatility index (PI) in the middle cerebral artery were measured with transcranial Doppler at an end-tidal CO2 of 4.5%. RESULTS: For sevoflurane Vmca was 18% lower [95% confidence interval (CI) 12-22%; P < 0.00001] and PI was 23% higher (95% CI 12-33%; P = 0.0013) than for isoflurane. Mean arterial blood pressure did not differ between the two agents. The minimum alveolar concentration (MAC) fraction necessary to reach the intended AAI level was 13% higher (95% CI 5-20%; P = 0.0079) with sevoflurane than with isoflurane. CONCLUSION: Sevoflurane induced less cerebral vasodilation than isoflurane at the same depth of anesthesia, measured by AAI, and hence seems more favorable for clinical neuroanesthesia. In our opinion the difference between sevoflurane and isoflurane in the MAC fraction required to attain the same AAI level demonstrates the limitations of MAC in defining the level of anesthesia.

Desflurane results in higher cerebral blood flow than sevoflurane or isoflurane at hypocapnia in pigs.

BACKGROUND: In clinical neuroanaesthesia, the increase in cerebral blood flow (CBF) and intracranial pressure caused by the cerebral vasodilative effects of an inhalational anaesthetic agent is counteracted by the cerebral vasoconstriction induced by hypocapnia. Desflurane and sevoflurane may have advantages over the more traditionally used isoflurane in neuroanaesthesia but their dose-dependent vasodilative effects at hypocapnia have not been compared in the same model using truly equipotent minimal alveolar concentrations (MACs). METHOD: Desflurane, sevoflurane and isoflurane were administered in a randomized order to six pigs at 0.5 and 1.0 MAC. The intra-arterial xenon clearance technique was used to calculate CBF. Blood pressure was invasively monitored. Cerebral and systemic physiological variables were recorded first at normocapnia (PaCO2 5.6 kPa) and then at hypocapnia (PaCO2 3.5 kPa). Electroencephalographic (EEG) activity was continuously recorded. RESULTS: None of the three agents abolished cerebrovascular reactivity to hyperventilation, and at 0.5 MAC all had similar effects on CBF at hypocapnia. Desflurane at 1.0 MAC was associated with 16% higher CBF (P = 0.027) at hypocapnia than isoflurane, and with 24% higher CBF (P = 0.020) than sevoflurane. There was no seizure activity in the EEG. CONCLUSION: More cerebral vasodilation at hypocapnia with high doses of desflurane than with sevoflurane or isoflurane indicates that desflurane might be less suitable for neuroanaesthesia than sevoflurane and isoflurane.

Cerebral blood flow at 0.5 and 1.0 minimal alveolar concentrations of desflurane or sevoflurane compared with isoflurane in normoventilated pigs.

Whether desflurane and sevoflurane have clinical advantages over isoflurane in neuroanesthesia is much debated. A porcine model was used for comparison of desflurane and sevoflurane with isoflurane with respect to their cerebrovascular effects. The minimal alveolar concentration (MAC) of each of the three agents was first determined in a standardized manner in six domestic juvenile pigs to enhance comparison reliability. Six other pigs were then anesthetized with isoflurane, desflurane, and sevoflurane, given in sequence to each pig in an even crosswise order with the first agent also used to maintain anesthesia during surgical preparation. Cerebral blood flow (CBF) was calculated from the clearance curve of intraarterially injected 133Xe. The mean arterial pressure (MAP) was invasively monitored. The estimated cerebrovascular resistance (CVRe) was calculated by dividing MAP with CBF, thereby approximating the cerebral perfusion pressure with MAP. For both MAC levels, the trend for CBF was desflurane > isoflurane > sevoflurane, and the trend for MAP and CVRe was sevoflurane > isoflurane > desflurane. Statistical comparison of desflurane and sevoflurane with isoflurane with respect to CBF and MAP revealed two statistically significant differences-namely, that CBF at 1.0 MAC desflurane was 17% higher than CBF at 1.0 MAC isoflurane (P =.0025) and that MAP at 1.0 MAC sevoflurane was 16% higher than MAP at 1.0 MAC isoflurane (P =.011). Consequently, in this study at normocapnia, these agents did not seem to differ much in their cerebral vasodilating effects at lower doses. At higher doses, however, desflurane, in contrast to sevoflurane, was found to induce more cerebral vasodilation than isoflurane.

PCR-generated linear DNA fragments utilized as a hantavirus DNA vaccine.

The field of DNA vaccines has grown rapidly, and since most such vaccines involve the inoculation of large circular DNA molecules previously propagated in bacteria, several inconveniences (e.g. the presence of antibiotic resistance genes, impurities from bacterial cultures or inefficient uptake of the large and bulky plasmid DNA molecules to the nucleus) are debated. In this study, we have explored the possibility of using smaller and more flexible PCR-generated linear DNA fragments instead. Phosphorothioate (PTO)-modified primers were used successfully to protect the PCR-generated DNA fragments from exonuclease degradation, and by using a nuclear localization signal-peptide to target the linear DNA to the nucleus the immune response against the encoded antigen was further improved. This approach was tested in cell culture using a sensitive reporter system and in vivo with DNA encoding the amino-terminus of the Puumala hantavirus nucleocapsid protein. Our results indicate that linear DNA fragments have a great potential as a genetic vaccine and phosphorothioate modification in combination with a nuclear localization signal peptide increase the stability and targets the linear DNA molecules to the nucleus resulting in an improved biological response examined both in vitro and in vivo.

Kinesin-dependent movement on microtubules precedes actin-based motility of vaccinia virus.

Vaccinia virus, a close relative of the causative agent of smallpox, exploits actin polymerization to enhance its cell-to-cell spread. We show that actin-based motility of vaccinia is initiated only at the plasma membrane and remains associated with it. There must therefore be another form of cytoplasmic viral transport, from the cell centre, where the virus replicates, to the periphery. Video analysis reveals that GFP-labelled intracellular enveloped virus particles (IEVs) move from their perinuclear site of assembly to the plasma membrane on microtubules. We show that the viral membrane protein A36R, which is essential for actin-based motility of vaccinia, is also involved in microtubule-mediated movement of IEVs. We further show that conventional kinesin is recruited to IEVs via the light chain TPR repeats and is required for microtubule-based motility of the virus. Vaccinia thus sequentially exploits the microtubule and actin cytoskeletons to enhance its cell-to-cell spread.

LcrV is a channel size-determining component of the Yop effector translocon of Yersinia.

Delivery of Yop effector proteins by pathogenic Yersinia across the eukaryotic cell membrane requires LcrV, YopB and YopD. These proteins were also required for channel formation in infected erythrocytes and, using different osmolytes, the contact-dependent haemolysis assay was used to study channel size. Channels associated with LcrV were around 3 nm, whereas the homologous PcrV protein of Pseudomonas aeruginosa induced channels of around 2 nm in diameter. In lipid bilayer membranes, purified LcrV and PcrV induced a stepwise conductance increase of 3 nS and 1 nS, respectively, in 1 M KCl. The regions important for channel size were localized to amino acids 127-195 of LcrV and to amino acids 106-173 of PcrV. The size of the channel correlated with the ability to translocate Yop effectors into host cells. We suggest that LcrV is a size-determining structural component of the Yop translocon.

Apically exposed, tight junction-associated beta1-integrins allow binding and YopE-mediated perturbation of epithelial barriers by wild-type Yersinia bacteria.

Using polarized epithelial cells, primarily MDCK-1, we assessed the mode of binding and effects on epithelial cell structure and permeability of Yersinia pseudotuberculosis yadA-deficient mutants. Initially, all bacteria except the invasin-deficient (inv) mutant adhered apically to the tight junction areas. These contact points of adjacent cells displayed beta1-integrins together with tight junction-associated ZO-1 and occludin proteins. Indeed, beta1-integrin expression was maximal in the tight junction area and then gradually decreased along the basolateral membranes. Wild-type bacteria also opened gradually the tight junction to paracellular permeation of different-sized markers, viz., 20-, 40-, and 70-kDa dextrans and 45-kDa ovalbumin, as well as to their own translocation between adjacent cells in intimate contact with beta1-integrins. The effects on the epithelial cells and their barrier properties could primarily be attributed to expression of the Yersinia outer membrane protein YopE, as the yopE mutant bound but caused no cytotoxicity. Moreover, the apical structure of filamentous actin (F-actin) was disturbed and tight junction-associated proteins (ZO-1 and occludin) were dispersed along the basolateral membranes. It is concluded that the Yersinia bacteria attach to beta1-integrins at tight junctions. Via this localized injection of YopE, they perturb the F-actin structure and distribution of proteins forming and regulating tight junctions. Thereby they promote paracellular translocation of bacteria and soluble compounds.

The V-antigen of Yersinia is surface exposed before target cell contact and involved in virulence protein translocation.

Type III-mediated translocation of Yop effectors is an essential virulence mechanism of pathogenic Yersinia. LcrV is the only protein secreted by the type III secretion system that induces protective immunity. LcrV also plays a significant role in the regulation of Yop expression and secretion. The role of LcrV in the virulence process has, however, remained elusive on account of its pleiotropic effects. Here, we show that anti-LcrV antibodies can block the delivery of Yop effectors into the target cell cytosol. This argues strongly for a critical role of LcrV in the Yop translocation process. Additional evidence supporting this role was obtained by genetic analysis. LcrV was found to be present on the bacterial surface before the establishment of bacteria target cell contact. These findings suggest that LcrV serves an important role in the initiation of the translocation process and provides one possible explanation for the mechanism of LcrV-induced protective immunity.

Yersinia outer proteins (YOPS) E, K and N are antigenic but non-protective compared to V antigen, in a murine model of bubonic plague.

The pathogenic Yersiniae produce a range of virulence proteins, encoded by a 70 kb plasmid, which are essential for infection, and also form part of a contact-dependent virulence mechanism. One of these proteins, V antigen, has been shown to confer a high level of protection against parenteral infection with Y. pestis in murine models, and is considered to be a protective antigen. In this study, the protective efficacy of V antigen has been compared in the same model with that of other proteins (YopE, YopK and YopN), which are part of the contact-dependent virulence mechanism. Mice immunised with two intraperitoneal doses of V antigen or each of the Yops, administered with either Alhydrogel or interleukin-12, produced high antigen-specific serum IgG titres. As shown in previous studies, V+Alhydrogel was fully protective, and 5/5 mice survived a subcutaneous challenge with 90 or 9x10(3) LD50's of Y. pestis GB. In addition, these preliminary studies also showed that V+IL-12 was partially protective: 4/5 or 3/5 mice survived a challenge with 90 or 9x10(3) LD50's, respectively. In contrast, none of the mice immunised with the Yops survived the challenges, and there was no significant delay in the mean time to death compared to mice receiving a control protein. These results show that using two different vaccine regimens, Yops E, K and N, failed to elicit protective immune responses in a murine model of plague, whereas under the same conditions, V antigen was fully or partially protective.

Functional conservation of the effector protein translocators PopB/YopB and PopD/YopD of Pseudomonas aeruginosa and Yersinia pseudotuberculosis.

Virulent Yersinia species cause systemic infections in rodents, and Y. pestis is highly pathogenic for humans. Pseudomonas aeruginosa, on the other hand, is an opportunistic pathogen, which normally infects only compromised individuals. Surprisingly, these pathogens both encode highly related contact-dependent secretion systems for the targeting of toxins into eukaryotic cells. In Yersinia, YopB and YopD direct the translocation of the secreted Yop effectors across the target cell membrane. In this study, we have analysed the function of the YopB and YopD homologues, PopB and PopD, encoded by P. aeruginosa. Expression of the pcrGVHpopBD operon in defined translocation-deficient mutants (yopB/yopD) of Yersinia resulted in complete complementation of the cell contact-dependent, YopE-induced cytotoxicity of Y. pseudotuberculosis on HeLa cells. We demonstrated that the complementation fully restored the ability of Y. pseudotuberculosis to translocate the effector molecules YopE and YopH into the HeLa cells. Similar to YopB, PopB induced a lytic effect on infected erythrocytes. The lytic activity induced by PopB could be prevented if the erythrocytes were infected in the presence of sugars larger than 3 nm in diameter, indicating that PopB induced a pore of similar size compared with that induced by YopB. Our findings show that the contact-dependent toxin-targeting mechanisms of Y. pseudotuberculosis and P. aeruginosa are conserved at the molecular level and that the translocator proteins are functionally interchangeable. Based on these similarities, we suggest that the translocation of toxins such as ExoS, ExoT and ExoU by P. aeruginosa across the eukaryotic cell membrane occurs via a pore induced by PopB.

YopK of Yersinia pseudotuberculosis controls translocation of Yop effectors across the eukaryotic cell membrane.

Introduction of anti-host factors into eukaryotic cells by extracellular bacteria is a strategy evolved by several Gram-negative pathogens. In these pathogens, the transport of virulence proteins across the bacterial membranes is governed by closely related type III secretion systems. For pathogenic Yersinia, the protein transport across the eukaryotic cell membrane occurs by a polarized mechanism requiring two secreted proteins, YopB and YopD. YopB was recently shown to induce the formation of a pore in the eukaryotic cell membrane, and through this pore, translocation of Yop effectors is believed to occur (Håkansson et al., 1996b). We have previously shown that YopK of Yersinia pseudotuberculosis is required for the development of a systemic infection in mice. Here, we have analysed the role of YopK in the virulence process in more detail. A yopK-mutant strain was found to induce a more rapid YopE-mediated cytotoxic response in HeLa cells as well as in MDCK-1 cells compared to the wild-type strain. We found that this was the result of a cell-contact-dependent increase in translocation of YopE into HeLa cells. In contrast, overexpression of YopK resulted in impaired translocation. In addition, we found that YopK also influenced the YopB-dependent lytic effect on sheep erythrocytes as well as on HeLa cells. A yopK-mutant strain showed a higher lytic activity and the induced pore was larger compared to the corresponding wild-type strain, whereas a strain overexpressing YopK reduced the lytic activity and the apparent pore size was smaller. The secreted YopK protein was found not to be translocated but, similar to YopB, localized to cell-associated bacteria during infection of HeLa cells. Based on these results, we propose a model where YopK controls the translocation of Yop effectors into eukaryotic cells.

Fibreoptic laryngotracheoscopy via the laryngeal mask airway in children.

BACKGROUND: Fibreoptic laryngotracheoscopy via the laryngeal mask airway-previously reported in adults but not in children-gives a better endoscopic view of the upper airway than does endoscopy via an endotracheal tube. METHOD: The endoscopic procedure was carried out via a size 2 laryngeal mask in 4 spontaneously breathing children between 1 and 6 years old. Anaesthesia was induced and maintained with halothane in 50-100% oxygen. RESULTS: The laryngeal mask was found to enable laryngotracheoscopy with a flexible 5.0-mm fibreoptic endoscope with no technical difficulties. Spontaneous ventilation could be readily preserved throughout the endoscopic procedure. CONCLUSIONS: In children anaesthetized with halothane, flexible fibreoptic laryngotracheoscopy via a laryngeal mask is a useful method-offering technical advantages not achieved otherwise-provided that generally approved restrictions to the use of a laryngeal mask airway are taken into account.

Cell-surface-bound Yersinia translocate the protein tyrosine phosphatase YopH by a polarized mechanism into the target cell.

YopH is translocated by cell-surface-bound bacteria through the plasma membrane to the cytosol of the HeLa cell. The transfer mechanism is contact dependent and polarizes the translocation to only occur at the contact zone between the bacterium and the target cell. More than 99% of the PTPase activity is associated with the HeLa cells. In contrast to the wild-type strain, the yopBD mutant cannot deliver YopH to the cytosol. Instead YopH is deposited in localized areas in the proximity of cell-associated bacteria. A yopN mutant secretes 40% of the total amount of YopH to the culture medium, suggesting a critical role of YopN in regulation of the polarized translocation. Evidence for a region in YopH important for its translocation through the plasma membrane of the target cell but not for secretion from the pathogen is provided.

Virulence plasmid-encoded YopK is essential for Yersinia pseudotuberculosis to cause systemic infection in mice.

The virulence plasmid common to pathogenic Yersinia species encodes a number of secreted proteins denoted Yops (Yersinia outer proteins). Here, we identify and characterize a novel plasmid-encoded virulence determinant of Yersinia pseudotuberculosis, YopK. The yopK gene was found to be conserved among the three pathogenic Yersinia species and to be homologous to the previously described yopQ and yopK genes of Y. enterocolitica and Y. pestis, respectively. Similar to the other Yops, YopK expression and secretion were shown to be regulated by temperature and by the extracellular Ca2+ concentration; thus, yopK is part of the yop regulon. In addition, YopK secretion was mediated by the specific Yop secretion system. In Y. pseudotuberculosis, YopK was shown neither to have a role in this bacterium's ability to resist phagocytosis by macrophages nor to cause cytotoxicity in HeLa cells. YopK was, however, shown to be required for the bacterium to cause a systemic infection in both intraperitoneally and orally infected mice. Characterization of the infection kinetics showed that, similarly to the wild-type strain, the yopK mutant strain colonized and persisted in the Peyer's patches of orally infected mice. A yopE mutant which is impaired in cytotoxicity and in antiphagocytosis was, however, found to be rapidly cleared from these lymphoid organs. Neither the yopK nor the yopE mutant strain could overcome the primary host defense and reach the spleen. This finding implies that YopK acts at a different level during the infections process than the antiphagocytic YopE cytotoxin does.