Antibiotic resistance is one of the most serious medical problems in the world. To counteract the increase in antibiotic resistance, new rapid and effective analytical methods are needed. To effectively treat infections in critically ill patients, optimal antibiotic dosages are required. DrugLog® is an instrument that uses a spectrophotometric method to analyze antibiotics in plasma in the wavelength range 200-800 nm. The aim of this study was to do a method validation of the instrument DrugLog®.     The study material that was used was whole blood from healthy donor and routine citrate plasma samples from the laboratory. The precision of the method and stability of plasma, the best way to filtrate lipids from plasma and four antibiotics (meropenem, cefotaxime, vancomycin, piperacillin/tazobactam) were investigated.     The precision of the method, measured as CV% was less than 0.62 and stability plasma showed a CV% of 135.74 after 24 h in room temperature. The stability for the different antibiotics after 24 h in room temperature showed a CV% of 8.11 for meropenem, 40.80 for vancomycin, 16.55 for cefotaxime and 2.92 for the combination antibiotic piperacillin/tazobactam. It was also determined that bacterial filter was the best way to remove lipids from plasma.     In conclusion DrugLog® is a suitable instrument to analyze concentration of antibiotics in patients during antibiotic treatment, however further validations are needed.

Contrast media are commonly used as an enhancement in several diagnostic imaging methods, which in today’s healthcare often are combined with blood works in diagnostics and surgical preparations, as well as to follow up on the patient’s recovery. To save time and money for both the hospital and the patients themselves, the ability to carry out both the radiological examination and the blood works within the same hospital visit would be preferred. However, there have been indications of a potential interference from the contrast media used, and therefore a waiting period is in place. The aim of this study was therefore to see if that waiting period was warranted by testing if contrast media does cause a significant interference in the most common analyses. This was investigated by infusing pooled samples with either iohexol or gadoteric acid, the active components of the most common contrast agents, at either a full dosage or a half dosage. These samples were then run by standard protocol and the results compared to control samples. The results showed that while some analyses proved affected, others proved unaffected or only insignificantly so. Some of the affected analyses were sodium, activated partial thrombin time and hemoglobin. While some analyses such as prostate specific antigen and prothrombin time were unaffected. Analysis of more samples is necessary to confirm the results, but the overall consensus is that while most analyses are unaffected the effects are too large and uncertain to comfortably disregard the waiting time.

Giardia intestinalis is the most common parasite found in dogs and cats. It is traditionally diagnosed using a microscope. These methods include direct immunofluorescence, DIF, and zinc sulphate centrifugal flotation, ZnSO4 C-flotation. However, there are commercially available SNAP tests such as the FASTest® GIARDIA strip that is often used by dogs and cats owner to detect Giardia. The aim of this study was to compare the sensitivity, cost and labor intensity of these three methods for detection of Giardia. To investigate this, 150 samples from dogs and cats were examined at the National Veterinary Institute in Sweden. The samples were a mixture of diarrheic and non-diarrheic stool. Of the 150 stool samples 100 samples were examined with FASTest® GIARDIA strip while 150 samples were examined with DIF and ZnSO4 C-flotation. The results indicated that FASTest® GIARDIA strip had a sensitivity of 66.18 %, a cost of 100 Swedish crowns (SEK) per sample and was the easiest test to use. ZnSO4 C-flotation had a sensitivity of 89.90 %, cost 418.75 SEK and took about 15 minutes to perform. DIF had 100 % sensitivity and specificity and due to that it was used as a standard reference method. The cost for DIF was 300 SEK and took more than an hour to perform per sample. The conclusion from this study is that, FASTest® GIARDIA strip is not a recommended test for detection of Giardia despite their low cost and easiness to use. DIF and ZnSO4 C-flotation remain a better diagnostic option for detection of Giardia.

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No description available.

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The antibiotic class β-lactams include drugs such as penicillins, cephalosporines, carbapenems and monobactams which mechanism of action is to inhibit cell-wall synthesis. Bacteria have developed several mechanisms to counter β-lactams. Bacteria can defend themselves from antibiotics by releasing enzymes that attack the antibiotic compound itself by hydrolysis, target alteration or redox reactions. Presence of antibiotics can also trigger a downregulation of genes coding for antibiotic binding proteins, as well as upregulation of proteins that serves as channel and pump proteins that ensure no accumulation of antibiotics occurs in the cytosol. The aim with the study was to investigate the presence of three plasmid-mediated genes (blaFOX, blaCIT(CMY-2) and blaMOX) coding for ampicillin resistance (pAmpC) in water isolates sampled from Helge River, Kristianstad. The detection of genes was done according to a previous optimized protocol for Real-Time PCR with SYBR™Green chemistry (duplex blaCIT(CMY-2)/blaMOX and singleplex blaFOX). The method proved not to be robust for multiplex PCR, only the singelplex for the gene blaFOX could produce valid results. 30 of 96 isolates were deemed as positive for the gene, whereas 27 of 79 were considered clinical relevant. Among the 27 isolates, 16 also harbored other genes for resistance (13 blaCTX-M, 2 blaOXA, 1 blaTEM and 1 blaSHV). One isolate carried on three resistancegenes (blaFOX, blaCTX-M och blaTEM). A majority of the positive isolates, 20 out of 27, were sampled near the pumpstation. The findings indicate that Helge river might be a reservoir for dissemination of antibiotic resistance genes.

Efter tappning av 450 mL helblod från frivilliga blodgivare separeras blodet till plasma, erytrocyter och trombocyter. Kontroller tas för att undersöka komponentframställningens resultat. I plasmaenheterna kontrolleras bland annat att antalet erytrocyter är under 6x109 per liter. Helblod innehåller normalt 4-6x1012 erytrocyter per liter. Massiv blödning är den främsta indikationen för plasmatransfusion. En transfusion är förknippad med risker såsom transfusion related acute lung injury (TRALI) och transfusion associated circulatory overload (TACO). Syftet med examensarbetet var att hitta en beslutsgräns för bestämning av erytrocytinnehållet i plasmakomponenter som framställs vid beredning av blodkomponenter inför transfusion. Gränsen skulle fastställas genom en jämförelse mellan teststickan Multistix 8 SG, Body fluid-programmet i hematologiinstrumentet Advia 2120 samt manuell räkning i Bürkers räknekammare. Analys skedde av 38 prover varav 18 prover fick tillsats av extra erytrocyter för att antingen överstiga kontrollgränsen eller finna övergången till stickans högsta nivå. Medelvärdet och medianen för de kvantitativa resultaten från Bürkers räknekammare för 20 godkända kontrollerna, fördelade efter teststickans kategorier, beräknades. Samtliga resultaten låg mellan 0,063x109/L och 2,08x109/L. Av de 38 prover som analyserades erhöll 37 ett resultat i form <10x109/L på Advia 2120. Utifrån de erhållna resultaten fastslogs gränsen vid vilken en komponentkontroll garanteras ett godkänt resultat till ≤2+ på teststickan. Vid resultat 3+ ska en konfirmerande kvantitativ analys utföras. Den slutsats som kunde dras var att teststickan Multistix 8 SG kan användas som en screeningmetod vid analys av erytrocytinnehållet i de tillverkade plasmakomponenterna. Slutsatsen blev även att det Adviaprogram som användes inte är lämpligt för analys av erytrocytinnehållet i plasma. / Following blood donation of 450 mL of whole blood from volunteer donors, the blood is separated into plasma, erythrocytes and thrombocytes. Controls are performed to investigate the separation performance. E.g. the plasma units are not allowed to contain more than 6x109 erythrocytes per liter. Whole blood does normally contain 4-6x1012 erythrocytes per liter. The primary indication for plasma transfusion is massive bleeding, a treatment mainly associated with risks such as transfusion related acute lung injury (TRALI) and transfusion associated circulatory overload (TACO). The aim of the thesis work was to find a decision limit for the determination of the erythrocyte content in plasma components produced prior to transfusion. The limit was to be determined by a comparison between the Multistix 8 SG test stick, Body fluid program in the Advia 2120 hematology instrument and manual count in the Bürker counting chamber. Analysis were performed on 38 samples, of which 18 samples were prepared by addition of extra erythrocytes to either exceed the control limit or find the transition point to the highest result level of the stick. The average and median of the quantitative results from the Bürker counting chamber for the 20 approved controls, broken down by the categories on the stick, were calculated. All results were between 0.063x109/L and 2.08x109/L. Of the 38 samples analyzed, 37 received a result <10x109/L on the Advia 2120. Based on these results, the decision limit at which a component control is guaranteed an approved result was determined to ≤2+ on the test stick. In the case of a 3+ result, a confirmatory quantitative analysis must be performed. The conclusion was that the test stick Multistix 8 SG could be used as a screening method for analyzing the erythrocyte content of the plasma components produced. The conclusion was also that the Adviaprogram used is not suitable for analysis of the erythrocyte content in plasma.

Slutsteget i immunförsvarets komplementaktivering innefattar en klyvning av komplementprotein C5, till C5a och C5b, vilket initierar bildandet av terminalt komplementkomplex (TCC) som i form av membran-attack-komplex (MAC) bildar cytotoxiska porer i bland annat gramnegativa bakterier. Bildandet av MAC kan blockeras av endogena regulatorer och TCC frisätts då som ett lösligt komplex, sC5b-9, i plasma. I examensarbetet studerades en variant av TCC, som i tidigare studier visats bildas oberoende av C3- och C5-konvertas när serum surgjorts till pH < 7,0 in vitro. Syftet med studien var att studera om detta atypiska TCC (aTCC) bildades hos grisar, som i en mekonium aspirationssyndrom (MAS)-modell, erhållit ett sänkt systemiskt pH in vivo. I syftet ingick också att etablera en ELISA-baserad metod för att analysera aTCC. I en sandwich ELISA användes monoklonal anti-C5a/C5a (desArg) (klon T13/9) som fångande antikropp och monoklonal anti-C9 (klon aE11) som detekterande antikropp för att analysera aTCC i plasmaprover från 18 MAS-grisar, samt i ett kontrollmaterial bestående av grisserum som surgjorts till pH 6,8 och 6,4 in vitro. Mängden aTCC i kontrollproverna ökade när pH sänktes men innehållet av aTCC i plasmaproverna minskade över MAS-studiens förlopp. När den relativa förändringen i aTCC relaterades till MAS-grisarnas slutliga pH kunde ett signifikant samband ses (p = 0,02) som visade att en större förändring i aTCC sammanföll med ett lägre slutligt pH. Nivåerna av aTCC var generellt sett högre i plasmaproverna jämfört med kontrollproverna vilket skulle kunnat bero på skillnader i plasma vs serum avseende aTCC eller att proverna kom från grisar med olika ålder och vikt. Avsaknad av grisspecifik standard och negativ kontroll samt lågt signal/brusförhållande bidrar till felkällor för analysen och denna kräver fortsatt optimering. / The late steps of complement activation involves a cleavage of complement protein C5, to C5a and C5b, which initiates the formation of terminal complement complex (TCC). The final complex is referred to as the membrane-attack-complex (MAC) which forms cytotoxic pores in, inter alia, gram-negative bacteria. The formation of MAC can be inhibited by endogenous regulators and the TCC is then released as a soluble complex, sC5b-9, in plasma. In the degree project, another type of TCC was studied, which in previous studies had shown to form independently of C3 and C5 convertases when serum was acidified to pH <7.0 in vitro. The purpose of the study was to investigate whether this atypical TCC (aTCC) was formed in piglets, which in a model of meconium aspiration syndrome (MAS), received a reduced systemic pH in vivo. The purpose was also to establish an ELISA for analyzing aTCC. Sandwich ELISA, with monoclonal anti-C5a / C5a (desArg) (clone T13/9) as a capture antibody and monoclonal anti-C9 (clone aE11) as a detection antibody, was used to analyse aTCC in plasma samples from 18 MAS piglets, and in control samples consisting of pig serum acidified to pH 6.8 and 6.4 in vitro. The amount of aTCC in the control samples increased when the pH was lowered, but the content of aTCC in the plasma samples decreased over the course of the MAS study. When the relative change in aTCC was related to the final pH of the MAS pigs, a significant relationship could be seen (p = 0.02) which showed that a major change in the aTCC coincided with a lower final pH. aTCC were generally higher in plasma samples compared with control samples, which could be due to differences in plasma vs serum for aTCC or that the samples came from pigs of different age and weight. Lack of pig-specific standard and negative control as well as low signal to noise ratio contribute to sources of error for the analysis and this requires continued optimization.

Marine bacteria play an important role in the marine nutrition cycles. About half of all sea-living bacteria can use light as an energy source, in addition to organic carbon compounds, which is important for survival as the seas becomes acidic and low in nutrition due to changing climate. The Flavobacteriaceae is a bacterial family that plays an important role in the degradation of complex organic compounds in natural environments. Dokdonia donghaensis MED134 belongs to the Flavobacteriaceae family and use both chemotrophy and phototrophy as a source of energy. For its phototrophic ability, the bacteria use proteorhodopsin, which is a membrane-bound proton pump. This allows the bacteria to survive and grow in nutrient-poor environments. The purpose of the study was to investigate with qPCR if there was a difference in gene expression for proteorhodopsin and isocitrate dehydrogenase in Dokdonia donghaensis MED134 depending on whether the bacteria had grown in constant light or darkness. Analysis with qPCR showed a significantly greater gene expression for proteorhodopsin in the bacteria grown in constant light for seven days, compared to those grown in constant light for three and four days and those grown in constant darkness. No significant difference could be demonstrated in gene expression for isocitrate dehydrogenase. This indicates that Dokdonia donghaensis MED134 in nutritional deficiency can use light as a source of energy for survival and further growth. By simulating climate change an adaptability could be seen through increased gene expression. Through this, further understanding of the role of bacteria in the marine ecosystem can be obtained and further research can be conducted as the marine climate issue becomes more relevant.