Interplay of Virulence Factors in Complement Resistance of Streptococcus pneumoniae

Streptococcus pneumoniae (pneumococcus) is a normal inhabitant of the human nasopharynx. Symptoms occur in only a small proportion of those who become carriers, but the ubiquity of the organism in the human population results in a large burden of disease. S. pneumoniae is the leading bacterial cause of pneumonia, sepsis, and meningitis worldwide, causing the death of a million children each year. Middle-ear infection is the most common clinical manifestation of mucosal pneumococcal infections. In invasive disease, S. pneumoniae gains access to the bloodstream and spreads to normally sterile parts of the body. The progression from asymptomatic colonization to disease depends on factors characteristic of specific pneumococcal strains as well as the status of host defenses. The polysaccharide capsule surrounding the bacterium is considered to be the most important factor affecting the virulence of pneumococci. It protects pneumococci from phagocytosis and also may determine its affinity to the respiratory epithelium. S. pneumoniae as a species comprises more than 90 different capsular serotypes, but not all of them are equally prevalent in human diseases. Invasive serotypes are rarely isolated from healthy carriers, but relatively often cause invasive disease. Serotypes that are carried asymptomatically for a long time behave like opportunistic pathogens, causing disease in patients who have impaired immune defenses. The complement system is a collection of blood and cell surface proteins that act as a major primary defense against invading microbes. Phagocytic cells with receptors for complement proteins can engulf and destroy pneumococcal cells opsonized with these proteins. S. pneumoniae has evolved a number of ways to subvert mechanisms of innate immunity, and this is likely to contribute to its pathogenicity. The capsular serotype, proteins essential for virulence, as well the genotype, may all influence the ability of pneumococcus to resist complement and its potential to cause disease. Immunization with conjugate vaccines produces opsonic antibodies, which enhance complement deposition and clearance of the bacteria. The pneumococcal vaccine included in the Finnish national immunization program in 2010 contains the most common serotypes causing invasive disease. Clinical data suggest that protection from middle-ear infection and possibly also from invasive disease depends largely on the capsular serotype, for reasons hitherto unknown. The general aim of this thesis is to assess the relative roles of the pneumococcal capsule and virulence proteins in complement evasion and subsequent opsonophagocytic killing. The main question is whether differences between serotypes to resist complement explain the different abilities of serotypes to cause disease. The importance of particular virulence factors to the complement resistance of a strain may vary depending on its genotype. Prior studies have evaluated the effect of the capsule and virulence proteins on complement resistance of S. pneumoniae by comparing only a few strains. In this thesis, the role of pneumococcal virulence factors in the complement resistance of the bacterium was studied in several genotypically different strains. The ability of pneumococci to inhibit deposition of the complement protein C3 on the bacterial surface was found to depend on the capsular serotype as well as on other features of the bacteria. The results suggest that pneumococcal histidine triad (Pht) proteins may play a role in complement inhibition, but their contribution depends on the bacterial genotype. The capsular serotype was found to influence complement resistance more than the bacterial genotype. A higher concentration of anticapsular antibodies was required for the opsonophagocytic killing of serotypes resistant to C3 deposition. The invasive serotypes were more resistant to C3 deposition than the opportunistic serotypes, suggesting that the former are better adapted to resist immune mechanisms controlling the development of invasive disease. The different susceptibilities of serotypes to complement deposition, opsonophagocytosis, and resultant antibody-mediated protection should be taken into account when guidelines for serological correlates for vaccine efficacy evaluations are made. The results of this thesis suggest that antibodies in higher quantity or quality are needed for efficient protection against the invasive serotypes.

High-yield bacterial expression and structural characterization of recombinant human insulin-like growth factor binding protein-2

The diverse biological activities of the insulin-like growth factors (IGF-1 and IGF-2) are mediated by the IGF-1 receptor (IGF-1R). These actions are modulated by a family of six IGF-binding proteins (ICFBP-1-6; 22-31 kDa) that via high affinity binding to the IGFs (K-D similar to 300-700 pM) both protect the IGFs in the circulation and attenuate IGF action by blocking their receptor access In recent years, IGFBPs have been implicated in a variety of cancers However, the structural basis of their interaction with IGFs and/or other proteins is not completely understood A critical challenge in the structural characterization of full-length IGFBPs has been the difficulty in expressing these proteins at levels suitable for NMR/X-ray crystallography analysis Here we describe the high-yield expression of full-length recombinant human IGFBP-2 (rhIGFBP-2) in Eschericha coli Using a single step purification protocol, rhIGFBP-2 was obtained with >95% purity and structurally characterized using NMR spectroscopy. The protein was found to exist as a monomer at the high concentrations required for structural studies and to exist in a single conformation exhibiting a unique intra-molecular disulfide-bonding pattern The protein retained full biologic activity. This study represents the first high-yield expression of wild-type recombinant human IGFBP-2 in E coli and first structural characterization of a full-length IGFBP (C) 2010 Elsevier Inc. All rights reserved

Improving outcome of childhood bacterial meningitis by simplified treatment : Experience from Angola

Background Acute bacterial meningitis (BM) continues to be an important cause of childhood mortality and morbidity, especially in developing countries. Prognostic scales and the identification of risk factors for adverse outcome both aid in assessing disease severity. New antimicrobial agents or adjunctive treatments - except for oral glycerol - have essentially failed to improve BM prognosis. A retrospective observational analysis found paracetamol beneficial in adult bacteraemic patients, and some experts recommend slow β-lactam infusion. We examined these treatments in a prospective, double-blind, placebo-controlled clinical trial. Patients and methods A retrospective analysis included 555 children treated for BM in 2004 in the infectious disease ward of the Paediatric Hospital of Luanda, Angola. Our prospective study randomised 723 children into four groups, to receive a combination of cefotaxime infusion or boluses every 6 hours for the first 24 hours and oral paracetamol or placebo for 48 hours. The primary endpoints were 1) death or severe neurological sequelae (SeNeSe), and 2) deafness. Results In the retrospective study, the mortality of children with blood transfusion was 23% (30 of 128) vs. without blood transfusion 39% (109 of 282; p=0.004). In the prospective study, 272 (38%) of the children died. Of those 451 surviving, 68 (15%) showed SeNeSe, and 12% (45 of 374) were deaf. Whereas no difference between treatment groups was observable in primary endpoints, the early mortality in the infusion-paracetamol group was lower, with the difference (Fisher s exact test) from the other groups at 24, 48, and 72 hours being significant (p=0.041, 0.0005, and 0.005, respectively). Prognostic factors for adverse outcomes were impaired consciousness, dyspnoea, seizures, delayed presentation, and absence of electricity at home (Simple Luanda Scale, SLS); the Bayesian Luanda Scale (BLS) also included abnormally low or high blood glucose. Conclusions New studies concerning the possible beneficial effect of blood transfusion, and concerning longer treatment with cefotaxime infusion and oral paracetamol, and a study to validate our simple prognostic scales are warranted.

Fast- and drift-ice communities in the Bothnian Bay and the impact of UVA radiation on the Baltic Sea ice ecology

The aim of this thesis was to study ecology of Baltic Sea ice from two perspectives. In the first two studies, sea-ice ecology from riverine-influenced fast ice to drift ice in the Bothnian Bay was investigated, whereas the last two studies focus on the sensitivity of sea-ice bacteria and algae to UVA examined in situ. The seasonal sea ice cover is one of the main characteristics of the Baltic Sea, and despite the brackish parental water, the ice structure is similar to polar ice with saline brine inclusions, the sea ice habitat. The decreasing seawater salinity from the northern Baltic Sea to the Bothnian Bay translates to decreasing brine volumes along the gradient, governing the size and community structure of the food webs in ice. However, the drift and fast ice in the Bothnian Bay may differ greatly in this sense, as drift ice may have been formed at more southern locations. Rafting and the formation of snow ice are common processes in the ice field of the Bothnian Bay. As evidenced in this thesis, rafting altered the vertical distribution of organisms and snow-ice formation provided habitable space in the better-illuminated, nitrogen-rich surface layer. The divergence between fast and drift ice became apparent at the more advanced stages, and chlorophyte biomass decreased from fast to drift ice, while the opposite held true for protozoan and metazoan biomass. The brine volumes affected the communities somewhat, and a higher percentage of flagellate species was generally linked to lower brine volumes, whereas chain-forming diatoms were mostly concentrated in layers with larger brine volumes. These results add to knowledge of the ecological significance of the ice cover lasting up to 7 months per year in this area. Sea-ice food webs are generally light-limited, but while increasing light irradiances typically enhance the primary production and further, the secondary production in sea ice, any increase in solar radiation also includes an increase in harmful UVA radiation. The Baltic Sea ice microbial communities were clearly sensitive to UVA and the responses were strongly linked to the earlier light history, as well as to the solar irradiances they were exposed to. The increased biomass of chlorophytes and pennate diatoms, when UVA was excluded, indicates that their normally minor contribution to the biomass in the upper layers of sea ice might be partly dictated by UVA. The effects of UVA on bacterial production in Baltic Sea ice mostly followed the responses in algal growth, but occasionally the exposure to UVA even enhanced the bacterial production. The dominant bacterial class, Flavobacteria, seemed to be UVA-tolerant, whereas all the Alpha-, Beta- and Gammaproteobacteria present in the surface layer showed UVA sensitivity. These results indicate that changes in the light field of ice may alter the community structure and affect the functioning of ice food webs, and are of importance when the effects of thinning of the ice cover are assessed.

Diversitet och tillväxtfrämjande egenskaper hos Stenotrophomonas-endofyter isolerade från Calliandra calothyrsus Meisn. rotknölar

The bacterial genus Stenotrophomonas comprises 12 species. They are widely found throughout the environment and particularly S. maltophilia, S. rhizophila and S. pavanii are closely associated with plants. Strains of the most common Stenotrophomonas species, S. maltophilia, promote plant growth and health, degrade natural and man-made pollutants and produce biomolecules of biotechnological and economical value. Many S. maltophilia –strains are also multidrug resistant and can act as opportunistic human pathogens. During an INCO-project (1998-2002) rhizobia were collected from root nodules of the tropical leguminous tree Calliandra calothyrsus Meisn. from several countries in Central America, Africa and New Caledonia. The strains were identified by the N2-group (Helsinki university) and some strains turned out to be members of the genus Stenotrophomonas. Several Stenotrophomonas strains induced white tumor- or nodule-like structures on Calliandra?s roots in plant experiments. The strains could, besides from root nodules, also be isolated from surface sterilized roots and stems. The purpose of my work was to investigate if the Stenotrophomonas strains i) belong to a new Stenotrophomonas species, ii) have the same origin, iii) if there are other differences than colony morphology between phase variations of the same strain, iv) have plant growth-promoting (PGP) activity or other advantageous effects on plants, and v) like rhizobia have ability to induce root nodule formation. The genetic diversity and clustering of the Stenotrophomonas strains were analyzed with AFLP fingerprinting to get indications about their geographical origin. Differences in enzymatic properties and ability to use different carbon and energy sources were tested between the two phases of each strain with commercial API tests for bacterial identification. The ability to infect root hairs and induce root nodule formation was investigated both using plant tests with the host plant Calliandra and PCR amplification of nodA and nodC genes for nodulation. The PGP activity of the strains was tested in vitro mainly with plate methods. The impact on growth, nitrogen content and nodulation in vivo was investigated through greenhouse experiments with the legumes Phaseolus vulgaris and Galega orientalis. Both the genetic and phenotypic diversity among the Stenotrophomonas strains was small, which proposes that they have the same origin. The strains brought about changes on the root hairs of Calliandra and they also increased the amount of root hairs. However, no root nodules were detected. The strains produced IAA, protease and lipase in vitro. They also showed plant a growth-promoting effect on G. orientalis, both alone and together with R. galegae HAMBI 540, and also activated nodulation among efficient rhizobia on P. vulgaris in greenhouse. It requires further research to get a better picture about the mechanisms behind the positive effects. The results in this thesis, however, confirm earlier studies concerning Stenotrophomonas positive impact on plants.

Campylobacter jejuni and C.coli in Finnish poultry production

Campylobacter, mainly Campylobacter jejuni and C. coli, are worldwide recognized as a major cause of bacterial food-borne gastroenteritis (World Health Organization 2010). Epidemiological studies have shown handling or eating of poultry to be significant risk factors for human infections. Campylobacter contamination can occur at all stages of a poultry meat production cycle. In summer 1999, every broiler flock from all three major Finnish poultry slaughterhouses was studied during a five month period. Caecal samples were taken in the slaughterhouses from five birds per flock. A total of 1 132 broiler flocks were tested and 33 (2.9%) of those were Campylobacter-positive. Thirty-one isolates were identified as C. jejuni and two isolates were C. coli. The isolates were serotyped for heat-stable antigens (HS) and genotyped by pulsed-field gel electrophoresis (PFGE). The most common serotypes found were HS 6,7, 12 and 4-complex. Using a combination of SmaI and KpnI patterns, 18 different PFGE types were identified. Thirty-five Finnish C. jejuni strains with five SmaI/SacII PFGE types selected among human and chicken isolates from 1997 and 1998 were used for comparison of their PFGE patterns, amplified fragment length polymorphism (AFLP) patterns, HaeIII ribotypes, and HS serotypes. The discriminatory power of PFGE, AFLP and ribotyping with HaeIII were shown to be at the same level for this selected set of strains, and these methods assigned the strains into the same groups. The PFGE and AFLP patterns within a genotype were highly similar, indicating genetic relatedness. An HS serotype was distributed among different genotypes, and different serotypes were identified within one genotype. From one turkey parent flock, the hatchery, six different commercial turkey farms (together 12 flocks) and from 11 stages at the slaughterhouse a total of 456 samples were collected during one and the half year. For the detection of Campylobacter both conventional culture and a PCR method were used. No Campylobacter were detected in either of the samples from the turkey parent flock or from the hatchery samples using the culture method. Instead PCR detected DNA of Campylobacter in five faecal samples from the turkey parent flock and in one fluff and an eggshell sample. Six out of 12 commercial turkey flocks were found negative at the farm level but only two of those were negative at slaughter. Campylobacter-positive samples within the flock at slaughter were detected between 0% and 94%, with evisceration and chilling water being the most critical stages for contamination. All of a total of 121 Campylobacter isolates were shown to be C. jejuni using a multiplex PCR assay. PFGE analysis of all isolates with KpnI restriction enzyme resulted in 11 PFGE types (I-XI) and flaA-SVR typing yielded nine flaA-SVR alleles. Three Campylobacter-positive turkey flocks were colonized by a limited number of Campylobacter genotypes both at the farm and slaughter level.In conclusion, in our first study in 1999 a low prevalence of Campylobacter in Finnish broiler flocks was detected and it has remained at a low level during the study period until the present. In the turkey meat production, we found that flocks which were negative at the farm became contaminated with Campylobacter at the slaughter process. These results suggest that proper and efficient cleaning and disinfection of slaughter and processing premises are needed to avoid cross-contamination. Prevention of colonization at the farm by a high level of biosecurity control and hygiene may be one of the most efficient ways to reduce the amount of Campylobacter-positive poultry meat in Finland. In Finland, with a persistent low level of Campylobacter-positive flocks, it could be speculated that the use of logistic slaughtering, according to Campylobacter status at farm, might have be advantageous in reducing Campylobacter contamination of retail poultry products. However, the significance of the domestic poultry meat for human campylobacteriosis in Finland should be evaluated.

The Biocontrol Agent Phlebiopsis gigantea: Efficacy and Impacts on the Stump Bacterial Biota and Conifer tree Defences

Phlebiopsis gigantea has been for a long time known as a strong competitor against Heterobasidion annosum and intensively applied as a biological control agent on stump surfaces of Picea abies in Fennoscandia. However, the mechanism underlying its antagonistic activity is still unknown. A primary concern is the possible impact of P. gigantea treatment on resident non-target microbial biota of conifer stumps. Additional risk factor is the potential of P. gigantea to acquire a necrotrophic habit through adaptation to living wood tissues. This study focused on the differential screening of several P. gigantea isolates from diverse geographical sources as well as the use of breeding approach to enhance the biocontrol efficacy against H. annosum infection. The results showed a significant positive correlation between growth rate in wood and high biocontrol efficacy. Furthermore, with aid of breeding approach, several progeny strains were obtained that had better growth rate and control efficacy than parental isolates. To address the issue of the potential of P. gigantea to acquire necrotrophic capability, a combination of histochemical, molecular and transcript profiling (454 sequencing) were used to investigate the interactions between these two fungi and ten year old P. sylvestris seedlings. The results revealed that both P. gigantea and H. annosum provoked strong necrotic lesions, but after prolonged incubation, P. gigantea lesions shrank and ceased to expand further. Tree seedlings pre-treated with P. gigantea further restricted H. annosum-induced necrosis and had elevated transcript levels of genes important for lignification, cell death regulation and jasmonic acid signalling. These suggest that induced localized resistance is a contributory factor for the biocontrol efficacy of P.gigantea, and it has a comparatively limited necrotrophic capability than H. annosum. Finally, to investigate the potential impact of P. gigantea on the stump bacterial biota, 16S rDNA isolated from tissue samples from stumps of P. abies after 1-, 6- and 13-year post treatment was sequenced using bar-coded 454 Titanium pyrosequencing. Proteobacteria were found to be the most abundant at the initial stages of stump decay but were selectively replaced by Acidobacteria at advanced stages of the decay. Moreover, P. gigantea treatment significantly decreased the bacterial richness at initial decay stage in the stumps. Over time, the bacterial community in the stumps gradually recovered and the negative effects of P. gigantea was attenuated.

Lower Gastrointestinal Microbiota in Health and Irritable Bowel Syndrome : Characterisation and Effect of Probiotic Intervention

The human gastrointestinal (GI) microbiota is a complex ecosystem that lives in symbiosis with its host. The growing awareness of the importance of the microbiota to the host as well as the development of culture-free laboratory techniques and computational methods has enormously expanded our knowledge of this microbial community. Irritable bowel syndrome (IBS) is a common functional bowel disorder affecting up to a fifth of the Western population. To date, IBS diagnosis has been based on GI symptoms and the exclusion of organic diseases. The GI microbiota has been found to be altered in this syndrome and probiotics can alleviate the symptoms, although clear links between the symptoms and the microbiota have not been demonstrated. The aim of the present work was to characterise IBS related alterations in the intestinal microbiota, their relation to IBS symptoms and their responsiveness to probiotic theraphy. In this thesis research, the healthy human microbiota was characterised by cloning and sequencing 16S rRNA genes from a faecal microbial community DNA pool that was first profiled and fractionated according to its guanine and cytosine content (%G+C). The most noticeable finding was that the high G+C Gram-positive bacteria (the phylum Actinobacteria) were more abundant compared to a corresponding library constructed from the unfractionated DNA pool sample. Previous molecular analyses of the gut microbiota have also shown comparatively low amounts of high G+C bacteria. Furthermore, the %G+C profiling approach was applied to a sample constructed of faecal DNA from diarrhea-predominant IBS (IBS-D) subjects. The phylogenetic microbial community comparison performed for healthy and IBS-D sequence libraries revealed that the IBS-D sample was rich in representatives of the phyla Firmicutes and Proteobacteria whereas Actinobacteria and Bacteroidetes were abundant in the healthy subjects. The family Lachnospiraceae within the Firmicutes was especially prevalent in the IBS-D sample. Moreover, associations of the GI microbiota with intestinal symptoms and the quality of life (QOL) were investigated, as well as the effect of probiotics on these factors. The microbial targets that were analysed with the quantitative real-time polymerase chain reaction (qPCR) in this study were phylotypes (species definition according to 16S rRNA gene sequence similarity) previously associated with either health or IBS. With a set of samples, the presence or abundance of a phylotype that had 94% 16S rRNA gene sequence similarity to Ruminococcus torques (R. torques 94%) was shown to be associated with the severity of IBS symptoms. The qPCR analyses for selected phylotypes were also applied to samples from a six-month probiotic intervention with a mixture of Lactobacillus rhamnosus GG, L. rhamnosus Lc705, Propionibacterium freudenreichii ssp. shermanii JS and Bifidobacterium breve Bb99. The intervention had been previously reported to alleviate IBS symptoms, but no associations with the analysed microbiota representatives were shown. However, with the phylotype-specific assays applied here, the abundance of the R. torques 94% -phylotype was shown to be lowered in the probiotic-receiving group during the probiotic supplementation, whereas a Clostridium thermosuccinogenes 85% phylotype, previously associated with a healthy microbiota, was found to be increased compared to the placebo group. To conclude, with the combination of methods applied, higher abundance of Actinobacteria was detected in the healthy gut than found in previous studies, and significant phylum-level microbiota alterations could be shown in IBS-D. Thus, the results of this study provide a detailed overview of the human GI microbiota in healthy subjects and in subjects with IBS. Furthermore, the IBS symptoms were linked to a particular clostridial phylotype, and probiotic supplementation was demonstrated to alter the GI microbiota towards a healthier state with regard to this and an additional bacterial phylotype. For the first time, distinct phylotype-level alterations in the microbiota were linked to IBS symptoms and shown to respond to probiotic therapy.

High-quality annotation of promoter regions for 913 bacterial genomes

Motivation: The number of bacterial genomes being sequenced is increasing very rapidly and hence, it is crucial to have procedures for rapid and reliable annotation of their functional elements such as promoter regions, which control the expression of each gene or each transcription unit of the genome. The present work addresses this requirement and presents a generic method applicable across organisms. Results: Relative stability of the DNA double helical sequences has been used to discriminate promoter regions from non-promoter regions. Based on the difference in stability between neighboring regions, an algorithm has been implemented to predict promoter regions on a large scale over 913 microbial genome sequences. The average free energy values for the promoter regions as well as their downstream regions are found to differ, depending on their GC content. Threshold values to identify promoter regions have been derived using sequences flanking a subset of translation start sites from all microbial genomes and then used to predict promoters over the complete genome sequences. An average recall value of 72% (which indicates the percentage of protein and RNA coding genes with predicted promoter regions assigned to them) and precision of 56% is achieved over the 913 microbial genome dataset.

Common epitopes of serum retinol-binding proteins: a study with monoclonal antibodies

Monoclonal antibodies were raised against purified chicken retinol-binding protein. These were characterised extensively with respect to their ability to recognize retinol-binding proteins from different species. The monoclonal antibodies exhibited differential recognition characteristics. Though the majority presented restricted reactivities, one out of the four monoclonal antibodies studied cross-reacted with retinol-binding proteins from all species tested so far.

The potential of microbial ecological indicators to guide ecosophisticated management of hydrocarbon-contaminated soils

Soil is an unrenewable natural resource under increasing anthropogenic pressure. One of the main threats to soils, compromising their ability to provide us with the goods and ecosystem services we expect, is pollution. Oil hydrocarbons are the most common soil contaminants, and they disturb not just the biota but also the physicochemical properties of soils. Indigenous soil micro-organisms respond rapidly to changes in the soil ecosystem, and are chronically in direct contact with the hydrophobic pollutants on the soil surfaces. Soil microbial variables could thus serve as an intrinsically relevant indicator of soil quality, to be used in the ecological risk assessment of contaminated and remediated soils. Two contrasting studies were designed to investigate soil microbial ecological responses to hydrocarbons, together with parallel changes in soil physicochemical and ecotoxicological properties. The aim was to identify quantitative or qualitative microbiological variables that would be practicable and broadly applicable for the assessment of the quality and restoration of oil-polluted soil. Soil bacteria commonly react on hydrocarbons as a beneficial substrate, which lead to a positive response in the classical microbiological soil quality indicators; negative impacts were accurately reflected only after severe contamination. Hydrocarbon contaminants become less bioavailable due to weathering processes, and their potentially toxic effects decrease faster than the total concentration. Indigenous hydrocarbon degrader bacteria, naturally present in any terrestrial environment, use specific mechanisms to improve access to the hydrocarbon molecules adsorbed on soil surfaces. Thus when contaminants are unavailable even to the specialised degraders, they should pose no hazard to other biota either. Change in the ratio of hydrocarbon degrader numbers to total microbes was detected to predictably indicate pollutant effects and bioavailability. Also bacterial diversity, a qualitative community characteristic, decreased as a response to hydrocarbons. Stabilisation of community evenness, and community structure that reflected clean reference soil, indicated community recovery. If long-term temporal monitoring is difficult and appropriate clean reference soil unavailable, such comparison could possibly be based on DNA-based community analysis, reflecting past+present, and RNA-based community analysis, showing exclusively present conditions. Microbial ecological indicators cannot replace chemical oil analyses, but they are theoretically relevant and operationally practicable additional tools for ecological risk assessment. As such, they can guide ecologically informed and sustainable ecosophisticated management of oil-contaminated lands.

Suolistopatogeenisten Escherichia coli -bakteerien esiintyvyys ripulipotilailla ja raa’oissa lihatuotteissa Burkina Fasossa sekä enterohemorraagisten E. coli -kantojen karakterisointi

Suolistopatogeeniset Escherichia coli -bakteerit eli ripulikolit aiheuttavat ihmisellä suolistoinfektioita. Kuten normaalimikrobiston E. coli -bakteerit, ne esiintyvät ihmisen lisäksi muiden nisäkkäiden, etenkin märehtijöiden, ja lintujen suolistossa. Lisäksi ne voivat esiintyä maaperässä ja vesistöissä. Ihminen voi saada tartunnan eläinperäisten elintarvikkeiden välityksellä tai juomalla eläinten tai ihmisen ulosteilla saastunutta vettä. Ripulikolit voidaan jakaa ainakin viiteen ryhmään perustuen niiden erilaisiin virulenssiominaisuuksiin: enteropatogeeninen E. coli (EPEC), enterotoksigeeninen E. coli (ETEC), enterohemorraaginen E. coli (EHEC), enteroinvasiivinen E. coli (EIEC) ja enteroaggregatiivinen E. coli (EAEC). EPEC aiheuttaa etenkin kehitysmaissa pikkulapsille ripulia. ETEC aiheuttaa turistiripulia ja vastasyntyneiden ripulia kehitysmaissa. EHEC aiheuttaa ripulia tai veriripulia, joka voi varsinkin pienillä lapsilla johtaa hemolyyttis-ureemiseen oireyhtymään (HUS) ja munuaisten vaurioitumiseen. EIEC aiheuttaa Shigellan kaltaista ripulia, joka voi olla veristä. EAEC on yhdistetty lähinnä pitkittyneisiin ripuleihin. Tutkimuksessa selvitettiin suolistopatogeenisten E. coli -bakteerien esiintyvyyttä Burkina Fasossa, josta ei ole saatavilla aikaisempaa tietoa ripulikolien esiintymisestä ihmisissä ja elintarvikkeissa. Ulostenäytteitä otettiin ripulia sairastavilta alle viisivuotiailta lapsilta maaseudulta kahdesta kylästä, Boromosta ja Gourcysta, ja maan pääkaupungista Ouagadougousta (110 näytettä). Lihanäytteitä (kanaa, nautaa, lammasta ja naudan suolta, jota käytetään ihmisravinnoksi) otettiin Ouagadougoun toreilla myytävistä kypsentämättömistä lihoista (120 näytettä). Näytteistä saadut bakteerisekaviljelmät tutkittiin monialukkeisella PCR-menetelmällä, joka tunnistaa viiden ripulikoliryhmän virulenssigeenejä. Lisäksi lihanäytteistä eristettiin 20 EHEC-kantaa shigatoksiinin stx-geenin havaitsemiseen perustuvalla pesäkehybridisaatiolla ja PCR-seulonnalla, ja karakterisoitiin mahdollisten virulenssiominaisuuksien selvittämiseksi. Tutkimus osoitti, että ripulikolien aiheuttamat suolistoinfektiot ovat yleisiä ripulia sairastavilla pikkulapsilla Burkina Fasossa. Ulostenäytteistä 59 % oli positiivisia. Useimmiten lapsilla esiintyi EAEC- (32 %) ETEC- (31 %) ja EPEC-patoryhmiä (20 %). EIEC- (2 %) ja EHEC-patoryhmiä (1 %) esiintyi vähän. Myös useamman patoryhmän sekainfektiot olivat yleisiä (24 %). Eri paikkakuntien välillä oli tilastollisesti merkitseviä eroja ripulikolien esiintymisessä. Gourcyssa ripulikoleja esiintyi useammin kuin Ouagadougoussa ja Boromossa. Tutkimuksessa kävi ilmi, että Ouagadougoun toreilla myytävissä lihoissa on paljon ripulikoleja. Lihanäytteistä 43 % oli positiivisia. Yleisimmin lihoissa esiintyi EHEC (28 %), EPEC (20 %), ETEC (8 %) ja EAEC (5 %). EIEC-ryhmää ei havaittu lihoissa. Myös useamman patoryhmän sekakontaminaatioita löytyi (17 %) lihoista. Ripulikolien esiintyvyydessä eri lihojen välillä ei ollut tilastollisesti merkitseviä eroja, kun tarkasteltiin kaikkia patoryhmiä yhdessä. Eri patoryhmien esiintyvyyttä tarkasteltaessa EHEC-patoryhmää ei esiintynyt ollenkaan kanassa ja ero oli tilastollisesti merkitsevä muihin lihoihin verrattuna. Lihoista eristetyt 20 EHEC-kantaa kuuluivat 14 eri serotyyppiin, joista osa on aikaisemmin eristetty suolistoinfektioihin ja HUSoireyhtymään sairastuneilta ihmisiltä. Kaikki kannat olivat stx1-positiivisia ja puolella oli lisäksi stx2-geeni, jota pidetään shigatoksiinin virulentimpana muotona. Kahdelta EHEC-kannalta löytyi myös ETECpatoryhmän lämpöstabiilin enterotoksiini Ia:n geeni eli kannat olivat kahden patoryhmän välimuotoa ja osoitus geenien siirtymisestä eri patoryhmien välillä. Vaikka nuorimmat näytteen antaneet lapsipotilaat tuskin söivät lihaa, sen voidaan ajatella silti olevan edustava näyte lasten elinympäristöstä, sillä lasten ruoka valmistetaan usein samoissa oloissa, joissa raakaa lihaa käsitellään. Saastunut liha voi siten olla pikkulasten ripulikoli-infektioiden aiheuttaja.

Kaupunkipurot Helsingissä - Veden laatu vuonna 2004

In recent years urban hydrology and individual urban streams have been in focus and subjects to research also in Helsinki. However, until now there has been lack of research covering simultaneously the whole area of the city of Helsinki. The aim of this study was to find out the general state of water quality in small urban streams in the city of Helsinki. 21 streams were studied: Mätäjoki, Korppaanoja, Mätäpuro, Näsinoja-Tuomarinkylänoja, Tuomarinkartanonpuro, Kumpulanpuro, Tapaninkylänpuro, Tapaninvainionpuro, Puistolanpuro, Longinoja, Säynäslahdenpuro, Viikinoja, Porolahdenpuro, Mustapuro, Marjaniemenpuro, Mellunkylänpuro, Vuosaarenpuro, Rastilanpuro, Ramsinkannaksenpuro, Skatanpuro and Yliskylänpuro. Water samples were collected from 48 sampling points, each stream having at least one point. Four water samples were collected from each point, sampling periods being 9.-11.2., 26.-28.4., 29.6.-1.7. and 25.-27.10.2004. Field measurements associated with water sampling included water temperature, oxygen concentration, pH and electrical conductivity. Water samples were analysed in the Laboratory of Physical Geography in the University of Helsinki and in the Environmental Laboratory of the City of Helsinki Environment Centre for following properties: suspended solids, dissolved substances, alkalinity, principal anions and cations (Na+, K+, Mg2+, Ca2+, F-, Cl-, NO3-, PO43- and SO42-), colour, turbidity, biological and chemical oxygen demand (BOD7 and CODMn-values), nutrient concentrations and bacterial indicators of hygienic quality. The main water quality issues found in this study were low oxygen levels in many streams and poor hygienic quality at least occasionally. E.g. in summer oxygen levels were under 60 % in every stream. Amount of total dissolved substances and nutrients were high in some of the streams studied. Compared to other Finnish streams the values of alkalinity and pH were higher. Although these problems were common, the variation between different streams and sampling points was significant. This was probably due to local conditions. Best overall water quality was found in Mätäpuro and Tuomarinkartanonpuro streams. Seasonal variation was evident in almost all water quality properties. For example the total amount of dissolved substances was largest in winter and decreased during the year. Colour and turbidity were smallest in winter and increased towards the end of the year. The same was true for suspended solids, which had smallest concentration in winter and greatest in autumn. It must be kept in mind that the spring samples were collected after the spring flood otherwise the largest suspended solid concentrations would have been expected in spring. Finnish general water quality classification was used to assess the quality of urban stream waters. Its suitability for small urban streams is not, however, completely trouble-free. This classification does not take into account the quick changes in such small streams but evaluates only the yearly mean values. This can oversimplify the picture of the water quality situation in the streams. Also in order to better reflect the urban environment the analysed water quality properties should also include total dissolved substances and e.g. concentrations of chloride and sodium.

Common Infrastructure for Finite-State Based Methods and Linguistics Descriptions

Finite-state methods have been adopted widely in computational morphology and related linguistic applications. To enable efficient development of finite-state based linguistic descriptions, these methods should be a freely available resource for academic language research and the language technology industry. The following needs can be identified: (i) a registry that maps the existing approaches, implementations and descriptions, (ii) managing the incompatibilities of the existing tools, (iii) increasing synergy and complementary functionality of the tools, (iv) persistent availability of the tools used to manipulate the archived descriptions, (v) an archive for free finite-state based tools and linguistic descriptions. Addressing these challenges contributes to building a common research infrastructure for advanced language technology.