Pharmacodynamic studies are important for the optimal use of antimicrobial agents. Combination antifungal therapy may be one method to improve outcome in invasive Candida infections. An in vitro kinetic model to study the pharmacodynamic effects of a combination of two antifungal agents with different elimination rates was developed and the pharmacodynamics of amphotericin B (AMB), voriconazole (VRC) or the combination was evaluated. Exposure to VRC inhibited the fungicidal activity of sequential doses of AMB against VRC-susceptible strains of C. albicans. The interaction was VRC dose-dependent. AMB activity was regained once VRC was removed or it increased gradually when the concentration of VRC had fallen below the minimum inhibitory concentration (MIC). The VRC-AMB interaction, however, was also present against strains of C. albicans, C. glabrata and C. krusei despite reduced VRC susceptibility. Against these strains the interaction was not predicted by the MIC value, suggesting that mechanisms of resistance may be of importance. Until more data are available, a reasonable recommendation is probably to avoid the sequential use of VRC followed by AMB and to use the combination of VRC and AMB for the treatment of Candida infections with caution. Only the unbound fraction of a drug is generally accepted as pharmacologically active. The activity of posaconazole (POS) with a protein binding of 98-99% was tested in serum against Candida species and compared with the calculated unbound serum concentration in protein-free media. Significant differences emerged at clinically relevant POS serum concentrations of 1.0 and 0.10 mg/l compared with the serum control regimen against one strain of C. lusitaniae. In RPMI 1640 the corresponding calculated unbound concentrations resulted in no effect for the low dose regimen compared with the RPMI 1640 control regimen. Further, against seven additional Candida strains tested, the effect of POS was greater in serum than in RPMI 1640. A flux from serum protein bound to fungal lanosterol 14α-demethylase bound POS may be the explanatory mechanism.

Long-term protective immunity depends on proper initiation of professional antigen-presenting cells (APCs). Autoimmune disorders and certain infections can cause disease through modulation of APCs and thereby affecting the outcome of these diseases. This work aimed to investigate the behaviour of different APC subsets during conditions known to cause improper immune responses. In Paper I, the effects of an anti-inflammatory compound called Rabeximod, intended for treatment of rheumatoid arthritis were investigated on different subsets of APCs. The results showed that Rabeximod affected the differentiation and behaviour of inflammatory subsets of dendritic cells (DCs) and macrophages while no effects were observed on anti-inflammatory subsets. Our findings suggest that Rabeximod acts by inhibiting the functionality of inflammatory subsets of APCs. In Paper II, the effects of different malaria derived stimuli such as hemozoin (Hz) and infected red-blood cells (iRBCs) on monocyte-derived dendritic cells (MoDCs) were investigated. Both stimuli triggered activation and migration of MoDCs. MoDCs exposed to iRBCs induced allogeneic T-cell proliferation while those exposed to Hz did not. These results indicate that different malaria derived stimuli may differently affect DCs and that this could lead to improper and inefficient T-cell activation. In Paper III, innate aspects of malarial immunity were compared in children from two sympatric ethnic groups. We observed decreased activation of APCs and severely supressed TLR responses in Dogon children as compared to Fulani. This may indicate an important role for TLR and APC activation in the Fulani, known to be better protected against malaria than the Dogon. In summary, detailed knowledge of APC activation will be helpful in the understanding of specific effector immune responses. This could in turn, improve treatment of inflammatory disorders as well as the generation of efficient vaccines against infectious diseases.

<p>Ribosomes are the universal ribonucleoprotein organelles that translate the genetic message from mRNA to protein. In prokaryotes, the ribosomal subunits are 30S and 50S subunit, which bind together during the translation process forming 70S ribosome. The ribosome is a highly dynamic structure, and acts as a working platform for the different factors involved in the process of converting the genetic information into protein.</p><p>Cryo-electron tomography (cryo-ET) is an emerging imaging technology that combines the potential of three-dimensional (3D) reconstruction at molecular resolution with a close-to-native preservation of the specimen. Here, we have applied this method to reconstruct rifampicin-treated <i>Escherichia coli</i> individual 30S subunits in vitro and in situ, and individual 50S subunits in situ. In the 30S subunit, the head, the platform and the body show large conformational movements relative to each other. The particles are grouped into three conformational groups according to the width/height ratios. Also, an S15 fusion protein derivative has been used as a physical reporter to localize S15 in the 30S subunit. In the 50S subunit, the L1 stalk, the L7/L12 stalk, the central protuberance (CP), and the peptidyl transferase center (PTC) cleft are the most dynamic and flexible parts in the reconstructed structures with clear movements indicated. Different locations of the tunnel in the central cross-sections through the in situ 50S subunits indicate a flexible pathway inside the large subunit. In addition, gross morphological changes were also been observed in our reconstructions. Our results demonstrate a considerable conformational flexibility among individual ribosomal subunits, both in vitro and in situ.</p><p>Translation is an essential process for all cells and organisms. Translation initiation is the rate-limiting step and the most highly regulated phase of translation process. Several regions along the mRNA have been reported to influence translation initiation. The Shine-Dalgarno (SD) sequence located 5-9 bases upstream of the initiation codon supports translation initiation by complementary binding to the Anti-Shine-Dalgarno (ASD) sequence on the 16S rRNA.</p><p>We have here compared how an SD⁺ sequence influences gene expression, if located upstream or downstream of an initiation codon. The positive effect of an upstream SD⁺ is confirmed. A downstream SD⁺ gives decreased gene expression. If an SD⁺ is placed between two potential initiation codons, initiation takes place predominantly at the second start site. The first start site is activated if the distance between this site and the downstream SD⁺ is enlarged and/or if the second start site is weakened. Upstream initiation is eliminated if a stable stem-loop structure is placed between this SD⁺ and the upstream start site. The results suggest that the two start sites compete for ribosomes that bind to an SD⁺ located between them. A minor positive contribution to upstream initiation resulting from 3’ to 5’ ribosomal diffusion along the mRNA is suggested. Since the location of SD⁺or SD-like sequences can strongly influence gene expression, this should be of significant evolutionary importance.</p>

<p>Idag lever cirka 33 miljoner människor med HIV/Aids. Totalt har omkring 30 miljoner människor avlidit sedan första Aids- fallet diagnostiserades år 1981. I västvärlden har HIV-smittade en tät kontakt med sjukvården eftersom de går på regelbundna kontroller, därmed är sjuksköterskor en yrkeskategori som träffar dessa patienter ofta. I utvecklingsländerna ser det ofta tyvärr annorlunda ut, HIV-smittade har ingen eller gles kontakt med sjukvården. Syftet med studien var att belysa sjuksköterskans upplevelser i omvårdnaden av HIV/Aids patienter. Uppsatsen genomfördes som en litteraturstudie där 14 vetenskapliga artiklar granskades, bearbetades och analyserades. Resultatet visade att sjuksköterskornas upplevelser av att vårda HIV/Aids patienter varierade. Positiva upplevelser kunde bland annat vara att arbetet var utvecklande och sjuksköterskorna kände sig viktiga för patienterna. Negativa upplevelser var bland annat rädsla och brist på resurser. Att vårda patienter med HIV/Aids kräver god kunskap om både sjukdomen och adekvata omvårdnadsåtgärder. Mer utbildning inom området efterlyses i sjuksköterskans grundutbildning. Förslag till vidare forskning är att undersöka vad sjuksköterskorna anser att man kunde göra för att ta bort de felaktiga negativa bilderna av sjukdomen och komma till rätta med de brister som finns idag.</p> / <p>Today about 33 million people have conflicted HIV/Aids. A total of about 30 million people have died of Aids since the first case was diagnosed in 1981. In the Industrial states people infected with HIV have close contact with medical care units, since they attend regular examinations. Due to this, nurses often come in contact with these patients. In the developing countries it is unfortunately not like that, HIV-infected has sparse or none contact with medical units. The purpose of this study was to illustrate nurses' experiences when caring for patients diagnosed with HIV/Aids. The study was carried out as a literature study where 14 scientific articles were reviewed, processed and analyzed. The result shows that nurses' experiences during their care of HIV/Aids patients varies. Among the positive experiences could be noticed that nurses felt that their work was important and was developing them. Among the negative experiences occurred fear and lack of resources. Medical care of patients with HIV/Aids demands good knowledge about the disease and adequate nursing care. Additional education is needed during nurses' basic education. A suggestion concerning further research includes researching what suggestions nurses have on how to correct faulty views of the disease and how to correct areas that need improvement.</p>

<p>Postoperativ sårinfektion är en komplikation som var tionde patient drabbas av efter ett kirurgiskt ingrepp. Det medför inte bara lidande och förlängd vårdtid för patienten utan kan även vara direkt livshotande. Den förlängda vårdtiden medför dessutom kostnader för samhället, och resurser skulle kunna användas till annan vård. Syftet med litteraturstudien var att beskriva sådana omvårdnadsåtgärder som sjuksköterskan kan vidta för att förebygga postoperativa sårinfektioner. Evidensbaserade omvårdnadsåtgärder som, var för sig minskar risken för att patienten ska drabbas av en postoperativ sårinfektion, och tillsammans utgör grunden för en säker vård. Databassökning av vetenskapligt material inom området ligger till grund för resultatet. Genom aktuell forskning presenteras och lyfts olika omvårdnadsåtgärder som reducerar risken för patienten att drabbas av en postoperativ sårinfektion. Viktiga omvårdnadsåtgärder är: korrekt hårborttagning, bibehållen normotermi, dusch med desinfektion, glukoskontroll, administrering av antibiotikaprofylax och postoperativ sårvård. Ny forskning inom området efterfrågas för att kunna följa utvecklingen, eftersom den befintliga är publicerad för många år sedan. För att patienten ska kunna erbjudas en säker vård behövs kontinuerlig utbildning under sjuksköterskeutbildningen men även i den kliniska verksamheten. Regelbunden uppföljning och utvärdering bör också ske i den kliniska verksamheten för att omvårdnadsåtgärderna ska vara effektiva.</p> / <p>Postoperative surgical wound infection is a complication that every tenth patient suffering after a surgical procedure. The consequences are the suffering and prolonged length of stay for the patient and can also be directly fatal. The prolonged duration of treatment is a high cost in society and resources could be used for other care. The purpose of literature review was to describe nurse’s interventions, which can be taken to prevent postoperative surgical wound infections. Evidence-based care interventions which reduce the risk of the patients suffering a postoperative surgical wound infection and together they represent a safe care. The result is based on search in databases for scientific materials in the subject area. Through current research highlights interventions which reduce the risk of the patient suffering a postoperative wound infection. Essential nursing interventions which are identified as: Hair removal, warming, shower with disinfectant, glucose monitoring, administration of antibiotic prophylaxis and wound care. New research in this area is requested to follow the developments since the current research is getting old. If the care should be safe for patient there must be education in nursing training as well as in the clinical work. Continuous follow-up should also occur in the clinical work in order to get feedback if the nursing interventions are effective.</p>

<p><strong>Abstract</strong></p><p><strong>Introduction</strong></p><p>Occurrence of bacteraemia and fungemia is a serious condition with high mortality and the incidence is increasing worldwide. The aim of this study was to survey the occurrence of bacteria and fungi in blood cultures from adult patients domiciled in the county of Gävleborg during one year and also to calculate the incidence and mortality in the same geographical area.</p><p><strong>Method</strong></p><p>This is a descriptive epidemiologic study, based on all episodes of blood cultures analyzed at the Microbiology laboratory, Gävle hospital during 2005. Patients from 20 years of age, domiciled in the county of Gävleborg at the date of drawing the blood culture, where included in the study. Criteria of exclusion were negative blood cultures and cultures which were classified as contaminants.</p><p><strong>Results</strong></p><p>Altogether there were 4 564 blood cultures analyzed, resulting in 524 (11 %) positive cultures for further study. There were 442 patients (48 % women) involved in 499 episodes with confirmed bacteraemia or fungemia. Gram positive bacteria represented 52 %, gram negative 45 % and fungi 3 %. The most frequently isolated bacterium was <em>Escherichia coli </em>followed by <em>Staphylococcus aureus. </em>In women, <em>Escherichia coli </em>was the most common bacterium, and there was a significant difference between the genders (<em>p= </em>0.004). In men, <em>Staphylococcus </em><em>aureus </em>was the dominant species (<em>p= </em>0.027<em>)</em>. <em>Streptococcus pneumoniae </em>was more common in women (<em>p= </em>0.005). The incidence of bacteraemia and fungemia in the county of Gävleborg was 235/100 000 inhabitants above the age of 20 (women, 223/100 000 men, 247/100 000). The incidence increased with age and the mean age was 70.2 years. The mortality within 30 days after the last positive blood culture was 22 % (97 patients). <em>Escherichia coli </em>was the most common bacteria diagnosed among those who died. The mortality in fungemia was 66 %. There was no significant difference in incidence or mortality between the two provinces Gästrikland and Hälsingland. Patients with bacteraemia and fungemia were initially cared for at all medical care units at the three hospitals in the county.</p><p><strong>Conclusion</strong></p><p>The incidence of bacteraemia/fungemia in the county of Gävleborg was 235/100 000 inhabitants. The most common bacteria in patients with confirmed bacteraemia were <em>Escherichia coli </em>and <em>Staphylococcus aureus. </em>Increasing age was a contributing risk factor. Patients with fungemia had considerably higher mortality compared to patients with bacteraemia. There where no significant differences in mortality between the two provinces.</p> / <p><strong>Introduktion</strong></p><p>Fynd av bakterier, bakteriemi, och svampar, fungemi, i blodbanan är ett allvarligt tillstånd med hög mortalitet och incidensen ökar i världen. Syftet med denna studie var att kartlägga vilka bakterier och svampar som förekom i alla blododlingar tagna under ett år från vuxna patienter i Gävleborgs län, samt att analysera incidens och mortalitet för bakteriemi och fungemi i länet.</p><p><strong>Metod</strong></p><p>Det är en deskriptiv epidemiologisk studie som utgår från analyserade blododlingar under år 2005 vid Enheten för Klinisk Mikrobiologi Laboratoriemedicin vid Gävle sjukhus. Till studien inkluderades personer från 20 års ålder som var mantalsskrivna i Gävleborgs län det datum som blododlingen utfördes. Exklusionskriterierna var negativa odlingssvar och svar som bedömdes som kontamination.</p><p><strong>Resultat</strong></p><p>Totalt analyserades 4 564 blododlingar, av vilka 524 (11 %) var positiva och bearbetades i denna studie. Det blev 442 patienter (48 % kvinnor) med 499 episoder av säkerställd bakteriemi eller fungemi. De grampositiva bakterierna stod för 52 %, gramnegativa bakterier 45 % och svampar 3 %. De enskilt vanligaste bakterierna var <em>Escherichia coli </em>och <em>Staphylococcus aureus. </em>För kvinnorna var <em>Escherichia coli </em>vanligast och det fanns en signifikant skillnad mellan könen (<em>p= </em>0,004 ), för männen var <em>Staphylococcus aureus </em>vanligast (<em>p= </em>0,027<em>)</em>. <em>Streptococcus pneumoniae </em>visade högre förekomst bland kvinnorna än männen (<em>p= </em>0,005). Incidensen för bakteriemi och fungemi i Gävleborgs län var 235/100 000 invånare äldre än 20 år (kvinnor, 223/100 000 och män, 247/100 000). Incidensen ökade med åldern och medelåldern var 70,2 år. Mortaliteten inom 30 dagar efter utförd blododling var 22 % (97 patienter). <em>Escherichia coli </em>var vanligast hos de avlidna. För patienter med fungemi var mortaliteten 66 %. Det påvisades ingen signifikant skillnad beträffande incidens eller mortalitet mellan länets båda landskap Gästrikland och Hälsingland. Patienter med bakteriemi och fungemi vårdades initialt på samtliga vårdenheter på länets tre sjukhus.</p><p><strong>Konklusion</strong></p><p>Incidensen för bakteriemi/fungemi i Gävleborgs län var 235/100 000 invånare. De vanligaste fynden vid säkerställd bakteriemi var <em>Escherichia coli </em>och <em>Staphylococcus aureus</em>. Ökande ålder var en riskfaktor. Patienter med fungemi hade avsevärt högre mortalitet än de med bakteriemi. Ingen skillnad påvisades mellan de två landskapen beträffande mortalitet.</p>

<p>Becoming antibiotic resistant is often associated with fitness costs for the resistant bacteria. This is seen as a loss of competitiveness against the antibiotic-sensitive wild-type in an antibiotic-free environment. In this study, the physiological alterations associated with fitness cost of antibiotic resistance <i>in vitro</i> (in the laboratory medium), and <i>in vivo</i> (in a mouse infection model), are identified in the model system of fusidic acid resistant (Fus^R) <i>Salmonella</i> <i>enterica</i> serovar Typhimurium.</p><p>Fus^R mutants have mutations in <i>fusA</i>, the gene that encodes translation elongation factor G (EF-G). Fus^R EF-G has a slow rate of regeneration of active EF-G·GTP off the ribosome, resulting in a slow rate of protein synthesis. The low fitness of Fus^R mutants <i>in vitro</i>, and <i>in vivo</i>, can be explained in part by a slow rate of protein synthesis and resulting slow growth. However, some Fus^R mutants with normal rates of protein synthesis still suffer from reduced fitness <i>in vivo</i>. We observed that Fus^R mutants have perturbed levels of the global regulatory molecule ppGpp. One consequence of this is an inefficient induction of RpoS, a regulator of general stress reponse and an important virulence factor for <i>Salmonella</i>. In addition, we found that Fus^R mutants have reduced amounts of heme, a co-factor of catalases and cytochromes. As a consequence of the heme defect, Fus^R mutants have a reduced ability to withstand oxidative stress and a low rate of aerobic respiration.</p><p>The pleiotropic phenotypes of Fus^R mutants suggest that antibiotic resistance can be associated with broad changes in bacterial physiology. Knowledge of physiological alterations that reduce the fitness of antibiotic-resistant mutants can be useful in identifying novel targets for antimicrobial agents. Drugs that alter the levels of global transcriptional regulators such as ppGpp or RpoS deserve attention as potential antimicrobial agents. Finally, the observation that Fus^R mutants have increased sensitivity to several unrelated classes of antibiotics suggests that the identification of physiological cost of resistance can help in optimizing treatment of resistant bacterial populations.</p>

<p>Antibiotic resistance is of current concern. Bacteria have become increasingly resistant to commonly used antibiotics and we are facing a growing resistance problem. The present thesis was aimed at studying the impact of antibiotic treatment on pathogenic bacteria as well as on the normal human microbiota, with focus on resistance development.</p><p>Among the factors that affect the appearance of acquired antibiotic resistance, the mutation frequency and biological cost of resistance are of special importance. Our work shows that the mutation frequency in clinical isolates of <i>Helicobacter pylori</i> was generally higher than for other studied bacteria such as <i>Enterobacteriaceae; </i>¼ of the isolates displayed a mutation frequency higher than<i> Enterobacteriaceae </i>defective<i> </i>mismatch repair mutants and could be regarded as mutator strains.</p><p>In <i>H. pylori</i>, clarithromycin resistance confers a biological cost, as measured by decreased competitive ability of the resistant mutants in mice. In clinical isolates, this cost could be reduced, consistent with compensatory evolution stabilizing the presence of the resistant phenotype in the population. Thus, compensation is a clinically relevant phenomenon that can occur in vivo.</p><p>Furthermore, our results show that clinical use of antibiotics selects for stable resistance in the human microbiota. This is important for several reasons. First, many commensals occasionally can cause severe disease, even though they are part of the normal microbiota. Therefore, stably resistant populations increase the risk of unsuccessful treatment of such infections. Second, resistance in the normal microbiota might contribute to increased resistance development among pathogens by interspecies transfer of resistant determinants.</p>

<p>In recent years molecular biology has become an integral part of the clinical laboratory. With an ever increasing number of methodologies and applications being presented each year it has increased our knowledge of how bacteria cause disease as well as our ability to predict disease outcome. </p><p>The main focus of this thesis has been to develop methods for identifying biomarkers and prediction methods for bacterial infectious diseases by taking advantage of the ever increasing possibilities of molecular biology. We applied cutting edge techniques in order to establish novel platforms for identifying and characterizing biomarkers of disease. </p><p>In paper one we describe a novel approach to measure levels of antibiotic resistance and viability of C. trachomatis, a method that is a clear improvement over existing techniques. In the second paper we describe the development of two assays designed to type pertussis toxin subunit 1 in circulating strains, in order to facilitate multi center studies for vaccine escape surveillance. In paper three we develop a novel microarray application designed to identify a large number of bacterial traits of H. pylori simultaneously with human genetic polymorphisms in order to identify a collection of risk factors that could be used as a prediction tool for gastric cancer risk. In the last paper we define the “antigenome” of H. pylori and identified 14 promising, previously unreported antigens as well as a number of potential biomarkers.</p><p>The platform technologies described in this collection of papers will hopefully help us identifying novel ways of fighting and predicting bacterial disease in future studies. </p>

Becoming antibiotic resistant is often associated with fitness costs for the resistant bacteria. This is seen as a loss of competitiveness against the antibiotic-sensitive wild-type in an antibiotic-free environment. In this study, the physiological alterations associated with fitness cost of antibiotic resistance in vitro (in the laboratory medium), and in vivo (in a mouse infection model), are identified in the model system of fusidic acid resistant (FusR) Salmonella enterica serovar Typhimurium. FusR mutants have mutations in fusA, the gene that encodes translation elongation factor G (EF-G). FusR EF-G has a slow rate of regeneration of active EF-G·GTP off the ribosome, resulting in a slow rate of protein synthesis. The low fitness of FusR mutants in vitro, and in vivo, can be explained in part by a slow rate of protein synthesis and resulting slow growth. However, some FusR mutants with normal rates of protein synthesis still suffer from reduced fitness in vivo. We observed that FusR mutants have perturbed levels of the global regulatory molecule ppGpp. One consequence of this is an inefficient induction of RpoS, a regulator of general stress reponse and an important virulence factor for Salmonella. In addition, we found that FusR mutants have reduced amounts of heme, a co-factor of catalases and cytochromes. As a consequence of the heme defect, FusR mutants have a reduced ability to withstand oxidative stress and a low rate of aerobic respiration. The pleiotropic phenotypes of FusR mutants suggest that antibiotic resistance can be associated with broad changes in bacterial physiology. Knowledge of physiological alterations that reduce the fitness of antibiotic-resistant mutants can be useful in identifying novel targets for antimicrobial agents. Drugs that alter the levels of global transcriptional regulators such as ppGpp or RpoS deserve attention as potential antimicrobial agents. Finally, the observation that FusR mutants have increased sensitivity to several unrelated classes of antibiotics suggests that the identification of physiological cost of resistance can help in optimizing treatment of resistant bacterial populations.