1. The response of microbes to anthropogenically induced perturbations in the Gulf of Trieste (northern Adriatic Sea) : dissertationNeža Orel, Tinkara Tinta, 2025, doktorska disertacija Opis: Marine ecosystems are constantly influenced by anthropogenic pressures, including inputs of nutrients, pollutants, and allochthonous microorganisms. These inputs can disrupt the dynamics of marine microbiomes, which encompass the majority of biomass and highest diversity of all life forms, controlling most biogeochemical cycles in the ocean realm. Despite their importance for public safety and potential influence on marine ecosystems functioning, knowledge of the impact of these perturbations on coastal microbiome dynamics remains in its infancy. The research objectives of this dissertation were to: (1) characterize the composition of the coastal microbiome in anthropogenically impacted coastal ecosystem; (2) investigate the genetic potential of selected microorganisms for pathogenicity and ecological adaptations; and (3) explore the effects of wastewater on the dynamics and functioning of the coastal microbiome, as well as its implications for the biogeochemical state of the ecosystem.
To address the first aim, we conducted a year-round in situ survey of the pelagic microbiome within anthropogenically impacted coastal ecosystem, focusing on the seasonal and spatial dynamics of traditional and alternative faecal bacterial indicators. We used a culture-independent approach combined with 16S rRNA amplicon sequencing, which overcome limitations of culture-based methods. This analysis revealed that the microbiome was primarily structured by seasonal changes, regardless of proximity to pollution sources. The statistical tool and oceanographic model we applied indicated that riverine water serves as a key vector for introducing allochthonous microbes. This study underscores the importance of molecular approaches combined with statistical and oceanographic modelling for advancing environmental health assessments and detecting microbial indicators.
To address the second aim, we applied whole-genome sequencing to assess the pathogenic potential and genomic features of selected Vibrio isolates. During the analysis, we identified cross-contamination in one isolate, which presented an opportunity to evaluate the effectiveness of bioinformatics workflows for contaminant removal and genome recovery. We reconstructed high-quality genomes of one Vibrio isolate from both axenic and contaminated cultures. Genomic analyses revealed that this isolate belongs to a sub-lineage of Vibrio campbellii associated with diseases in marine organisms. Moreover, this genome harboured a novel Vibrio plasmid linked to bacterial defense mechanisms and horizontal gene transfer, potentially offering a competitive advantage to this putative pathogen. This study highlights the utility of WGS and advanced bioinformatics in overcoming challenges posed by non-axenic cultures and provides new insights into the genomic characteristics of V. campbellii.
To address the third aim, we conducted a short-term microcosm experiment simulating wastewater discharge into coastal seawater, testing two types of wastewaters: (a) unfiltered, containing nutrients, pollutants, and allochthonous microbes, and (b) pre-filtered, retaining only nutrients and pollutants. Our results showed that wastewater, significantly increased nutrient levels (dissolved organic carbon, ammonium, orthophosphate). Using a multi-omics approach with measurements of microbial metabolic activity, we found that nutrient enrichment significantly influenced bacterial metabolism. This was evidenced by enriched protein profiles and increased leucine aminopeptidase and olease activity, indicating bacterial degradation of complex proteins and lipids. At the same time, the phosphate input resulted in a decreased alkaline phosphatase activity, with important implications for phosphorus cycling. Overall, wastewater primarily induced functional shifts in coastal microbiomes, highlighting the resilience and functional redundancy of coastal microbial communities and hence the biogeochemical processes they operate.
Ključne besede: coastal microbiome, anthropogenic impact, wastewater pollution, bacterial community dynamics, allochthonous microorganisms, potential pathogens, Vibrio campbellii, multi-omics, whole-genome assembly, dissertations
Objavljeno v RUNG: 16.04.2025; Ogledov: 210; Prenosov: 2
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2. DISTRIBUTION OF ENTERIC VIRUSES IN THE GULF OF TRIESTE AND THEIR INTERACTIONS WITH ENVIRONMENTAL AND BIOLOGICAL PARAMETERSJosé Manuel Carita Gonçalves, 2018, doktorska disertacija Opis: The available classical diagnostic methods, due to many disadvantages, do not allow effective detection of pathogenic enteric viruses in environmental samples. Due to low concentrations of pathogenic viruses in the sea, it is important to develop an effective concentration procedure for their successful detection.
In the first part of the doctoral thesis, we focused primarily on the development of a protocol for an effective concentration of pathogenic enteric viruses in coastal water samples. Monolithic chromatographic columns (BIAseparations) were used for the concentration of rotaviruses and noroviruses, prior to the detection with reverse transcription quantitative PCR in real time (RT-qPCR). We tested the efficiency of concentration using columns of various chemical properties and selected pathogenic enteric viruses (rotavirus and norovirus). Among them, hydrophobic interaction monolithic column (CIM® C4) was the most effective. CIM C4 was used to optimize the concentration step and tested in waters with different salinities.
The presence of concentrated viruses was confirmed by RT-qPCR and transmission electron microscope. We have developed a protocol that enables rapid concentration of viruses in coastal waters of various salinities and can be used on-site.
The presence of RoV and NoV was surveyed, using the developed concentration protocol, prior to one-step RT-qPCR molecular detection, in the inner part of the Bay of Koper, in mussel farming areas and a swimming area. Rotaviruses, noroviruses and fecal indicator bacteria were frequently detected in the inner part of the Bay of Koper.
Rotaviruses and noroviruses were detected in the studied area, with higher rates close to the outfall of the wastewater treatment plant in the estuary of river Rižana and were also detected in the middle of the Bay of Koper and in areas used for recreation and mussel farming.
The results show that water bodies, which are otherwise defined as suitable for bathing or mussel farming, based on the results of fecal indicator bacteria, still contain low concentrations of pathogenic enteric viruses.
In addition to human pathogenic enteric viruses and faecal coliforms, changes in abundance of bacteria and virus particles were studied in relation to temperature, salinity, inorganic and organic nutrient concentrations in the organically polluted Rižana estuary. Preliminary results showed spatially and seasonally changes in bacterial and viral particles abundance, and bacterial composition spatially and seasonally. However, seasonality plays a greater role in bacterial dynamics.
Ključne besede: Concentration of viruses, Enteric viruses, Rotavirus, Norovirus, Feacal coliforms, Feacal contamination, qPCR, RT-qPCR, Monolithic columns, Sewage, Seasonal dynamics, Concentration, Coastal environment, Gulf of Trieste
Objavljeno v RUNG: 02.07.2018; Ogledov: 6555; Prenosov: 214
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3. Odziv mikrobne združbe na mešanico nevarnih spojinNeža Orel, 2017, magistrsko delo Opis: Morske bakterije so najbolj številčni organizmi v morskem ekosistemu in imajo pomembno vlogo v biogeokemičnih procesih kroženja snovi. Preko različnih točkovnih in netočkovnih virov v morsko okolje vstopajo onesnažila, ki predstavljajo tveganje za zdravje ljudi, morskih organizmov in povzročajo spremembe ekosistema. Na podlagi raznolike občutljivosti mikroorganizmov na onesnaženje nam odziv bakterijske združbe lahko služi kot pokazatelj stanja v okolju. V tej študiji je bil opravljen laboratorijski eksperiment v zaprtem sistemu, z namenom preverjanja interakcij med onesnažili in mikroorganizmi v morski vodi Tržaškega zaliva. Naravna mikrobna združba je bila izpostavljena dvema mešanicama kemikalij v koncentracijah, ki jih Evropska zakonodaja predpisuje kot varne ter mešanici z 10-krat povišanimi koncentracijami. S pomočjo mikrobioloških analiz za določanje bakterijskega števila, hitrosti rasti bakterijske populacije ter z analizami bakterijske združbe smo želeli ugotoviti kolikšen je njihov vpliv na naravno mikrobno združbo v dveh različnih obdobjih (poleti, spomladi). Rezultati kažejo, da je dodatek onesnažil omogočil hitro rast bakterij in porast v številu. Bakterijsko število je bilo nižje samo v vzorcih izpostavljenih onesnažilom pri nižjih koncentracijah predpisanih mejnih vrednosti kot v kontrolnem vzorcu morske vode. V vzorcih izpostavljenih onesnažilom smo zaznali predvsem povišano rast bakterij predstavnikov družine Alteromonadaceae, medtem ko so bili nekateri predstavniki bakterij družine Rhodobacteraceae inhibirani. Rezultati so potrdili potencialno uporabo mikrobioloških analiz za zaznavanje sprememb, ki jih povzroči izpostavljenost naravne bakterijske združbe mešanici nevarnih kemikalij, ter opozorili na vprašanje dopustnih mejnih vrednosti predpisanih koncentracij onesnažil.
Ključne besede: Onesnaženje morja, toksičnost, mikrobiologija morja, bakterijska združba, prednostna lista nevarnih kemikalij
Objavljeno v RUNG: 07.12.2017; Ogledov: 8922; Prenosov: 266
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4. Metode določanja mikroorganizmov prisotnih v balastnih vodahTina Govednik, 2015, diplomsko delo Opis: Tujerodne vrste predstavljajo svetovni problem v morskih in sladkovodnih okoljih ter posledično ustvarjajo nevarnost za zdravje ljudi in okolja ter morsko biotsko raznovrstnost. Čezoceanske plovbe in promet omogočajo globalne povezave in istočasno prenos patogenih in drugih organizmov. Glavno pot vnosa tujerodnih vrst v okolje predstavlja balastna voda in sediment v ladijskih tankih. Z obstoječimi tehnologijami in priporočenimi standardi, ki jih je predlagala Mednarodna pomorska organizacija (IMO) lahko delno obvladujemo biološko onesnaževanje. Do sedaj nobena tehnologija ni bila dovolj učinkovita, da bi lahko odstranila vse organizme iz balastnih vod. Zato je bila predlagana študija določanja biološkega in kemičnega stanja v pristaniščih in s tem določanja kakovosti voda pred črpanjem v balastne tanke. V diplomski nalogi sem povzela nekatere predpise in razpoložljive tehnologije za določanje kakovosti in dezinfekcijsko obdelavo balastnih voda. Namen diplomske naloge je bil predvsem določitev koncentracij fekalnih koliformnih bakterij in celokupne združbe bakterij v ustju reke Rižane, kjer je glavni pomol za privez ladij v luki Koper, dveh drugih bazenov v neposredni bližini in na referenčni postaji na sredini Koprskega zaliva. Analize fekalnih koliformnih bakterij smo opravili z metodo membranske filtracije in pri tem uporabili različna gojišča, priporočena za analize koliformnih bakterij. Rezultati vzorcev morske vode kažejo, da je vzorčeno mesto v ustju reke Rižane onesnaženo, kamor se sicer izlivajo tudi odpadne vode čistilne naprave Koper. Pri vzorcih sedimenta so mikrobiološki rezultati zelo različni, vendar smo visoke vrednosti bakterij zaznali na vseh vzorčnih mestih v področju luke Koper. Primerjali smo tudi razlike v vrstni sestavi celokupne bakterijske združbe med vzorci vode in sedimenta različnih vzorčnih mest v Koprskem zalivu z metodo denaturacijske gradientne gelske elektroforeze.
Ključne besede: balastne vode, metode membranske filtracije, fekalni koliformi
Objavljeno v RUNG: 22.04.2015; Ogledov: 7285; Prenosov: 320
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5. Mikrobiološka aktivnost v obdobju pojavljanja polisaharidno bogatih makroagregatov v Tržaškem zalivu : diplomsko deloMeta Kalin, 2005, diplomsko delo Opis: V spomladanskem obdobju leta 2004 smo raziskovali razporeditev združbe heterotrofnih bakterij v vzhodnem delu Tržaškega zaliva. Bakterijske celice smo šteli pod epifluorescentnm mikroskopom. Bakterijsko produkcijo smo merili z metodo asimilacije radioaktivno označene spojine 3H-timidin in 3H-leucin. Število bakterij na dveh postajah Tržaškega zaliva se je spreminjalo od 0,2z109 do 2,1z109 celic/L. Najbolj izrazit padec bakterijske biomase smo beležili v maju, visoko biomaso pa smo beležili v času pojava sluzastih makroagregatov. Vzorce sluzastih makroagregatov smo pregledovali s pomočjo različnih mikroskopskih tehnik (svetlobni, epifluorescentni in elektronski mikroskop). V obdobju pojava sluzi smo izolirali posamezne bakterijske kolonije in jim določili osnovne morfološke znake. Z metodo fluorescentne in situ hibridizacije (FISH) smo opravili preliminarne analize določanje bakterijskih filogenetskih skupin. Bakterijske kolonije smo uporabili v dveh poskusih obogatitve morske vode z anorganskimi in organskimi hranili. Organsko hranilo je predstavljala sluz z visokim deležem polisaharidov. Odziv bakterijske združbe na dodatek hranil je bil hiter in se kaže v povečanju števila bakterijskih celic in visoki bakterijski produkciji že v prvih 24 urah. Sluz je heterotrofnim bakterijam služila kot vir energije in ogljika. V ta namen smo sledili koncentracijam celokupnih ogljikovih hidratov in proteinov. Spremembe v koncentraciji celokupnih ogljikovih hidratov so najverjetneje posledica povečanja bakterijske encimatske aktivnosti in istočasnega ponovnega privzema sproščenih ogljikovih hidratov. Preliminarni rezultati kažejo na velik pomen bakterijske heterotrofne združbe v procesih pretvorbe organske snovi v morskem ekosistemu.
Ključne besede: morje, mikrobiologija, kultura
Objavljeno v RUNG: 19.02.2015; Ogledov: 6067; Prenosov: 270
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8. Bacterial community structure and function in the Gulf of Trieste with some application studiesTinkara Tinta, 2011, doktorska disertacija Ključne besede: morske bakterije, bakterijske združbe, sezonska dinamika, DGGE, 16S rRNA klonske knjižnice, makroagregati, meduze, množično pojavljanje, Tržaški zaliv, disertacije
Objavljeno v RUNG: 15.10.2013; Ogledov: 7776; Prenosov: 459
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10. Ocena vpliva vnosa rečnih in komunalnih vod v Tržaški zalivTjaša Leban, 2008, diplomsko delo Ključne besede: diplomske naloge, odpadne vode, efluenti, indikatorski mikroorganizmi, fekalne bakterije, onesnaževalci, onesnaevanje, morje, obalne vode
Objavljeno v RUNG: 15.10.2013; Ogledov: 6952; Prenosov: 486
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