Molecular identification and localization of filamentous symbiotic bacteria associated with the hydrothermal vent annelid Alvinella pompejana

Alvinella pompejana is a polychaetous annelid that inhabits high temperature environments associated with active deep-sea hydrothermal vents along the East Pacific Rise. A unique and diverse epibiotic microflora with a prominent filamentous morphotype is found associated with the worm's dorsal integument. A previous study established the taxonomic positions of two epsilon proteobacterial phylotypes, 13B and 5A, which dominated a clone library of 16S rRNA genes amplified by PCR from the epibiotic microbial community of an A. pompejana specimen. In the present study deoxyoligonucleotide PCR primers specific for phylotypes 13B and 5A were used to demonstrate that these phylotypes are regular features of the bacterial community associated with A. pompejana. Assaying of other surfaces around colonies of A. pompejana revealed that phylotypes 13B and 5A are not restricted to A. pompejana. Phylotype 13B occurs on the exterior surfaces of other invertebrate genera and rock surfaces, and phylotype 5A occurs on a congener, Alvinella caudata. The 13B and 5A phylotypes were identified and localized on A. pompejana by in situ hybridization, demonstrating that these two phylotypes are, in fact, the prominent filamentous bacteria on the dorsal integument of A. pompejana. These findings indicate that the filamentous bacterial symbionts of A. pompejana are epsilon Proteobacteria which do not have an obligate requirement for A. pompejana.

Some observations on the absence of bacteria from acid waters in northwest England

This brief report concentrates on the effect of low pH on the initial stages of decomposition and the conditioning of incoming particulate carbon or detritus by microbes, particularly certain genera of filamentous bacteria. Although many previous reports have concentrated on bacterial decomposition processes, little attention has been given to the composition of the bacterial community and the role of its component parts, particularly in nutrient-poor waters which are provided with sources of organic carbon and reducing power in the form of poor quality detritus.

The in situ physiology of "Nostocoida limicola" II, a filamentous bacterial morphotype in bulking activated sludge, using fluorescence in situ hybridization and microautoradiography

The in situ physiology of the actinobacterial bulking and foaming filamentous bacterium "Nostocoida limicola" II was studied by fluorescence in situ hybridization/microautoradiography. Substrate assimilation patterns of pure cultures of this bacterium were different to those seen in activated sludge biomass samples. There was no evidence to suggest that "N. limicola" II preferred hydrophobic substrates, but evidence was produced to support the view that it is metabolically active under anaerobic conditions in activated sludge.

Diversity and identification of heterotrophic bacteria from Antarctic Rocks of the McMurdo Dry Valleys (Ross Desert)

Diversity of endolithic Dry Valley rock microorganisms was studied by evaluating the presence of morphotypes in enrichments. Storage of rock samples for 16 h over dry ice affected the diversity of endolithic organisms, especially that of algae and fungi. Diversity in various samples depended on rock location and exposure, on the rock type, and to some extent on the pH of the pulverized rock samples. In most cases sandstone contained more morphotypes than dolerite or granite. Presence of many different phototrophs resulted in greater diversity of the heterotrophs in the enrichments. Samples from Linnaeus Terrace and Battleship Promontory had higher morphotype (MT) numbers than those from more exposed sites such as New Mountain, University Valley, Dais, or Mt. Fleming. Beacon sandstone (13 samples) from Linnaeus Terrace varied greatly with respect to MT numbers, although the pH values ranged only from 4.2-5.3. The highest MT number of 24 per sample was obtained from the upper surface of a flat boulder tilted to the North. Only two MT's were found in a hard sandstone sample from the wind-exposed and more shaded east side of the Terrace. 15 sandstone samples from Battleship Promontory contained more diverse populations: there occurred a total of 131 different MT's in these samples as compared to only 68 in Linnaeus Terrace samples. Cysts of colorless flagellates were found in some Battleship Promontory samples; rnost samples were populated with a wealth of different cyanobacteria. Studies on the distribution of actinomycete morphotypes in Linnaeus Terrace sandstone revealed great differences between individual boulders. Identification tests and lipid analyses made with representative strains of the isolated 1500 pure cultures led to genus names such as Caulobacter, Blastobacter, Hyphomicrobium, Micrococcus, Arthrobacter, Brevibacterium, Corynebacterium, Bifidobacterium, Mycobacterium, Nocardia (Amycolata), Micromonospora, Streptomyces, Blastococcus, and Deinococcus. Our data demonstrate the great diversity of Antarctic endolithic microbial populations.

Meganema perideroedes gen. nov., sp nov., a filamentous alphaproteobacterium from activated sludge

An industrial wastewater treatment plant at Grindsted, Denmark, has suffered from bulking problems for several years caused by filamentous bacteria. Five strains were isolated from the sludge by micromanipulation, Phylogenetic analysis of the 16S rRNA gene sequences showed that the strains formed a monophyletic cluster in the Alphaproteobacteria, and they were phenotypically different from their closest relatives and from all hitherto known filamentous bacteria described (closest relative Brevundimonas vesicularis ATCC 11426(T), 89(.)8% sequence similarity). In pure culture, the cells (1(.)5-2(.)0 mu m) in filaments are Gram-negative and contain polyphosphate and polyhydroxyalkanoates. The optimum temperature for growth is 30 degrees C and the strains grow in 2 % NaCl and are oxidase- and catalase-positive. Ubiquinone 10 is the major quinone. The major fatty acid (C-18: 1 omega 7c) and smaller amounts of unsaturated fatty acids, 3-hydroxy fatty acids with a chain length of 16 and 18 carbon atoms and small amounts of 10-methyl-branched fatty acids with 18 carbon atoms (C-19: 0 10-methyl) affiliated the strains with the Methylobacterium/Xanthobacter group in the Alphaproteobacteria. The G + C content of the DNA is 42(.)9 mol% (for strain Gr1(T)). The two most dissimilar isolates by 16S rRNA gene comparison (Gr1(T) and Gr10; 97(.)7 % identical) showed 71(.)5 % DNA-DNA relatedness. Oligonucleotide probes specific for the pure cultures were designed for fluorescence in situ hybridization and demonstrated that two filamentous morphotypes were present in the Grindsted wastewater treatment plant. It is proposed that the isolates represent a new genus and species, Meganema perideroedes gen. nov., sp. nov. The type strain of Meganema perideroedes is strain Gr1(T) (=DSM 15528(T) =ATCC BAA-740(T)).

Microbial life and deposits in paper machine circuits

Väitöskirjani käsittele mikrobien ja erilaisten kemikaalien rooleja saostumien ja biofilmien muodostumisessa paperi- ja kartonkikoneilla. "Saostuma" tässä työssä tarkoittaa kiinteän aineen kertymää konepinnoille tai rajapinnoille konekierroissa, jotka on tarkoitettu massasulppujen, lietteiden, vesien tai ilman kuljetukseen. Saostumasta tulee "biofilmi" silloin kun sen oleellinen rakennekomponentti on mikrobisolut tai niiden tuotteet. Väitöstyöni työhypoteesina oli, että i. tietämys saostumien koostumuksesta, sekä ii. niiden rakenteesta, biologisista, fysikaalis-kemiallisista ja teknisistä ominaisuuksista ohjaavat tutkijaa löytämään ympäristöä säästäviä keinoja estää epätoivottujen saostumien muodostus tai purkaa jo muodostuneita saostumia. Selvittääkseni saostumien koostumista ja rakennetta käytin monia erilaisia analytiikan työkaluja, kuten elektronimikroskopiaa, konfokaali-laser mikroskopiaa (CLSM), energiadispersiivistä röntgenanalyysiä (EDX), pyrolyysi kaasukromatografiaa yhdistettynä massaspektrometriaan (Py-GCMS), joninvaihtokromatografiaa, kaasukromatografiaa ja mikrobiologisia analyysejä. Osallistuin aktiivisesti innovatiivisen, valon takaisinsirontaan perustuvan sensorin kehittämistyöhön, käytettäväksi biofilmin kasvun mittaukseen suoraan koneen vesikierroista ja säiliöistä. Työni osoitti, että monet paperinvalmistuksessa käytetyistä kemikaaleista reagoivat keskenään tuottaen orgaanisia tahmakerroksia konekiertojen teräspinnoille. Löysin myös kerrostumia, jotka valomikroskooppisessa tarkastelussa oli tulkittu mikrobeiksi, mutta jotka elektronimikroskopia paljasti alunasta syntyneiksi, alumiinihydroksidiksi joka saostui pH:ssa 6,8 kiertokuitua käyttävän koneen viiravesistä. Monet paperintekijät käyttävät vieläkin alunaa kiinnitysaineena vaikka prosessiolot ovat muuttuneet happamista neutraaleiksi. Sitä pidetään paperitekijän "aspiriinina", mutta väitöstutkimukseni osoitti sen riskit. Löysin myös orgaanisia saostumia, joiden alkuperä oli aineiden, kuten pihkan, saippuoituminen (kalsium saippuat) niin että muodostui tahmankasvua ylläpitävä alusta monilla paperi- ja kartonkikoneilla. Näin solumuodoiltaan Deinococcus geothermalista muistuttavia bakteereita kasvamassa lujasti teräskoepalojen pintaan kiinnittyneinä pesäkkeinä, kun koepaloja upotettiin paperikoneiden vesikiertoihin. Nämä deinokokkimaiset pesäkkeet voivat toimia jalustana, tarttumisalustana muiden mikrobien massoille, joka selittäisi miksi saostumat yleisesti sisältävät deinokokkeja pienenä, muttei koskaan pääasiallisena rakenneosana. Kun paperikoneiden käyttämien vesien (raakavedet, lämminvesi, biologisesti puhdistettu jätevesi) laatua tutkitaan, mittausmenetelmällä on suuri merkitys. Koepalan upotusmenetelmällä todettu biofilmikasvu ja viljelmenetelmällä mitattu bakteerisaastuneisuus korreloivat toisiinsa huonosti etenkin silloin kun likaantumisessa oli mukana rihmamaiseti kasvavia bakteereja. Huoli ympäristöstä on pakottanut paperi- ja kartonkikoneiden vesikiertojen sulkemiseen. Vesien kierrätys ja prosessivesien uudelleenkäyttö nostavat prosessilämpötilaa ja lisäävät koneella kiertävien kolloidisten ja liuenneiden aineiden määriä. Tutkin kiertovesien pitoisuuksia kolmessa eriasteisesti suljetussa tehtaassa, joiden päästöt olivat 0 m3, 0,5 m3 ja 4 m3 jätevettä tuotetonnia kohden, perustuen puhdistetun jäteveden uudelleen käyttöön. Nollapäästöisellä tehtaalla kiertovesiin kertyi paljon orgaanisesti sidottua hiiltä (> 10 g L-1), etenkin haihtuvina happoina (maito-, etikka-, propioni- ja voi-). Myös sulfaatteja, klorideja, natriumia ja kalsiumia kertyi paljon, > 1 g L-1 kutakin. Pääosa (>40%) kaikista bakteereista oli 16S rRNA geenisekvenssianalyysien tulosten perusteella sukua, joskin etäistä (< 96%) ainoastaan Enterococcus cecorum bakteerille. 4 m3 päästävältä tehtaalta löytyi lisäksi Bacillus thermoamylovorans ja Bacillus coagulans. Tehtaiden saostumat sisälsivät arkkeja suurina pitoisuuksina, ≥ 108 g-1, mutta tunnistukseen riittävää sekvenssisamanlaisuutta löytyi vain yhteen arkkisukuun, Methanothrix. Tutkimustulokset osoittivat että tehtaan vesikiertojen sulkeminen vähensi rajusti mikrobiston monimuotoisuutta, muttei estänyt liuenneen aineen ja kiintoaineen mineralisoitumista.

Food sources, behaviour, and distribution of hydrothermal vent shrimps at the Mid-Atlantic Ridge

Five species of bresilioid shrimp were investigated at seven hydrothermal sites on the Mid-Atlantic Ridge: Menez Gwen, Lucky Strike, Rainbow, Broken Spur, TAG, Snake Pit and Logatchev. Samples were prepared for analysis of stable isotopes, elemental composition and lipids. Shrimp behaviour was observed from the submersible ‘Alvin’ and in the laboratory aboard RV ‘Atlantis’. The distribution and zonation of the shrimp species was recorded. Juvenile shrimp of all species arrive at the vents carrying reserves of photosynthetic origin, built-up in the pelagic larval stages. These reserves are used while the shrimp metamorphose to the adult form and, in Rimicaris exoculata and Chorocaris chacei, while they develop epibiotic bacteria supporting structures, the modified mouthparts and the inside of the carapace. The main food of adult R. exoculata is filamentous bacteria that grow on these structures. The intermediate sizes of C. chacei also feed on such bacteria, but the final stage gets some food by scavenging or predation. Mirocaris species scavenge diverse sources; they are not trophically dependent on either R. exoculata or mussels. Adults of Alvinocaris markensis are predators of other vent animals, including R. exoculata. The dense swarms of R. exoculata, with their exosymbionts, can be compared to endosymbiont-containing animals such as Bathymodiolus and the vestimentiferan tube-worms of the Pacific vents. Such associations, whether endo- or ectosymbiotic, may be necessary for the development of flourishing communities at hydrothermal vents.

Bacterial symbiosis in Syssitomya pourtalesiana Oliver, 2012 (Galeommatoidea: Montacutidae), a bivalve commensal with the deep-sea echinoid Pourtalesia.

For the first time, bacterial symbiosis is recognized in the bivalve family Montacutidae of the superfamily Galeommatoidea. The ctenidial filaments of Syssitomya pourtalesiana Oliver, 2012 are extended abfrontally and a dense layer of bacteriocyte cells cover the entire surface behind a narrow ciliated frontal zone. The bacteria are extracellular and held within a matrix of epithelial extensions and microvilli. There is no cuticular layer (glycocalyx) covering the bacteria as in many thyasirid symbioses. The bacteriocytes hold more than one morphotype of bacteria, but bacilli, 1–3 μm in length, dominate. Scanning electron microscopy observations show a surface mat of filamentous bacteria over the extreme abfrontal surfaces. Filter feeding was confirmed by the presence of food particles in the stomach and the bivalve is presumed to be mixotrophic. Syssitomya is commensal and lives attached to the anal spines of the deep-sea echinoid Pourtalesia. In this position, echinoid feeding currents and echinoid faecal material may supply the bacteria with a variety of nutrient materials including dissolved organic matter.

Mauve Stingers (Pelagia noctiluca) as carriers of the bacterial fish pathogen Tenacibaculum maritimum

Aggregations or blooms of jellyfish are increasingly problematic for the aquaculture industry. Jellyfishassociated mass mortalities of sea-caged fish are most often caused by swarms of oceanic species like Pelagia noctiluca. These relatively large jellyfish get carried by tides and currents onto fish cages, causing them to break up into pathogenic nematocyst-containing pieces that are capable of passing through the mesh of the cages. The main effect on fish is gill damage leading to respiratory distress, but the lesions may also be compounded by bacterial infection, Tenacibaculum maritimum being one of the pathogens involved. In our previous study, we highlighted the ability of the jellyfish Phialella quadrata to carry this important pathogen. However, since these small jellyfish were collected around sea-cages of infected salmon, it was not possible to determine if the jellyfish or the fish themselves were the original source of the bacteria. Results of the current study demonstrate that these filamentous bacteria are present on the mouth of P. noctiluca that had no previous contact with farmed fish. These new results highlight the fact that some Cnidarian species harbour T. maritimum and suggest that jellyfishmight be a natural host for these bacteria whose environmental reservoir has not yet been determined.

Coprolites from the Cretaceous Bearpaw Formation of Saskatchewan

Coprolites from the Beechy Member of the Cretaceous Bearpaw Formation, southern Saskatchewan, presumably deposited by one or more species of mosasaur or large fish/shark, were recovered and analyzed using SEM/EDS. The data reveal the presence of pseudomorphous coccoid bacteria, potential filamentous bacteria, bacterial endospores and filamentous fungi. No recorded fossil plant or bone material could be identified, either within the highly compressed coprolitic mat-flattened full coprolite bolus - of recovered marine sediment encased in a mixed mat of hematite-apatite primary minerals heavily coated with Ca-smectite and nontronite, or the full coprolite bolus. The presence of fossil bacteria with morphological characteristics similar to those of endospores in other environments suggests that only robust microbial forms such as these survive diagenesis, partly with some carbon still intact, the remainder replaced with silica and iron. The data support the view that coprolites can serve as a useful source of information on the ancient microbial world. © 2012 Elsevier Ltd.

Reassessment of the Microbial Role in Mn-Fe Nodule Genesis in Andean Paleosols

The presence of Mn-Fe nodules in the epipedons (surface horizons) of paleosols of presumed Upper Neogene age in the northwestern Venezuelan Andes have been interpreted as products of inorganic oxidation and reduction processes operating over the full range of glacial and interglacial cycles that affected paleosol morphogenesis. New microscopic/chemical data from combined SEM-EDS-FIB analyses of representative Mn-Fe nodules indicate microbes play an important role in Mn/Fe precipitation leading to their genesis in alpine Mollisols (Argiustolls). Although the prevailing new data are based mainly on fossil forms of filamentous bacteria and fungi and other biogenic pseudomorphs that may represent the former resident bacteria, the presence of extant microbes must await field experiments/collection, followed by a molecular microbiology approach to determine the biological drivers of metal precipitation. As in other terrestrial niche environments, microbes are seen here to play a role, perhaps a key one, in the morphogenesis of paleosols of importance in upper Neogene paleoenvironmental reconstruction.

Investigation of the regulation mechanisms for bioplastics production from industrial residues

Dissertação para obtenção do Grau de Mestre em Biotecnologia

A dynamic knowledge-based decision support system to handle solids separation problems in activated sludge systems: development and validation

El sistema de fangs activats és el tractament biològic més àmpliament utilitzat arreu del món per la depuració d'aigües residuals. El seu funcionament depèn de la correcta operació tant del reactor biològic com del decantador secundari. Quan la fase de sedimentació no es realitza correctament, la biomassa no decantada s'escapa amb l'efluent causant un impacte sobre el medi receptor. Els problemes de separació de sòlids, són actualment una de les principals causes d'ineficiència en l'operació dels sistemes de fangs activats arreu del món. Inclouen: bulking filamentós, bulking viscós, escumes biològiques, creixement dispers, flòcul pin-point i desnitrificació incontrolada. L'origen dels problemes de separació generalment es troba en un desequilibri entre les principals comunitats de microorganismes implicades en la sedimentació de la biomassa: els bacteris formadors de flòcul i els bacteris filamentosos. Degut a aquest origen microbiològic, la seva identificació i control no és una tasca fàcil pels caps de planta. Els Sistemes de Suport a la Presa de Decisions basats en el coneixement (KBDSS) són un grup d'eines informàtiques caracteritzades per la seva capacitat de representar coneixement heurístic i tractar grans quantitats de dades. L'objectiu de la present tesi és el desenvolupament i validació d'un KBDSS específicament dissenyat per donar suport als caps de planta en el control dels problemes de separació de sòlids d'orígen microbiològic en els sistemes de fangs activats. Per aconseguir aquest objectiu principal, el KBDSS ha de presentar les següents característiques: (1) la implementació del sistema ha de ser viable i realista per garantir el seu correcte funcionament; (2) el raonament del sistema ha de ser dinàmic i evolutiu per adaptar-se a les necessitats del domini al qual es vol aplicar i (3) el raonament del sistema ha de ser intel·ligent. En primer lloc, a fi de garantir la viabilitat del sistema, s'ha realitzat un estudi a petita escala (Catalunya) que ha permès determinar tant les variables més utilitzades per a la diagnosi i monitorització dels problemes i els mètodes de control més viables, com la detecció de les principals limitacions que el sistema hauria de resoldre. Els resultats d'anteriors aplicacions han demostrat que la principal limitació en el desenvolupament de KBDSSs és l'estructura de la base de coneixement (KB), on es representa tot el coneixement adquirit sobre el domini, juntament amb els processos de raonament a seguir. En el nostre cas, tenint en compte la dinàmica del domini, aquestes limitacions es podrien veure incrementades si aquest disseny no fos òptim. En aquest sentit, s'ha proposat el Domino Model com a eina per dissenyar conceptualment el sistema. Finalment, segons el darrer objectiu referent al seguiment d'un raonament intel·ligent, l'ús d'un Sistema Expert (basat en coneixement expert) i l'ús d'un Sistema de Raonament Basat en Casos (basat en l'experiència) han estat integrats com els principals sistemes intel·ligents encarregats de dur a terme el raonament del KBDSS. Als capítols 5 i 6 respectivament, es presenten el desenvolupament del Sistema Expert dinàmic (ES) i del Sistema de Raonament Basat en Casos temporal, anomenat Sistema de Raonament Basat en Episodis (EBRS). A continuació, al capítol 7, es presenten detalls de la implementació del sistema global (KBDSS) en l'entorn G2. Seguidament, al capítol 8, es mostren els resultats obtinguts durant els 11 mesos de validació del sistema, on aspectes com la precisió, capacitat i utilitat del sistema han estat validats tant experimentalment (prèviament a la implementació) com a partir de la seva implementació real a l'EDAR de Girona. Finalment, al capítol 9 s'enumeren les principals conclusions derivades de la present tesi.

Potencial de aplicação do processo foto-fenton/solar como pré-tratamento de efluente da indústria de laticínios

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)