Ensaios de reprodução da medusa Phyllorhiza punctata para fins ornamentais

Ao longo das últimas décadas verificou-se um aumento do mercado de organismos ornamentais, tornando-se num mercado bastante lucrativo. Os cnidários são um dos grupos com maior interesse e valorização dentro dos organismos ornamentais, dentro destes as medusas têm captado a atenção como organismos ornamentais, apresentando uma grande margem de progressão, sendo atualmente consideradas como uma espécie emergente na indústria da aquariofilia. Embora existam muitas espécies de medusa com potencial ornamental, a seleção de uma espécie alvo para o uso em aquários domésticos obriga a que sejam eleitos alguns critérios de adequabilidade, nomeadamente não ser letal ou perigosa para o Homem. A espécie Phyllorhiza punctata é uma das espécies com muita procura no mercado ornamental. Este trabalho teve como objetivo estudar as condições ideais para obter medusas a partir da fase de pólipo. Neste sentido foram realizados vários ensaios de forma a otimizar cada fase do processo. Durante todo o processo de produção de medusas é necessário retirar os pólipos de um lugar para outro, colocando-os em novos substratos e esperar que se fixem. Neste sentido é importante que os pólipos se fixem aos substratos, para que ao serem manipulados não se soltem. Assim, foi realizado um estudo relativo à fixação de pólipos em diferentes substratos, nomeadamente: placa de 6 poços, caixa de petri, gobelet de vidro e de plástico. Verificou-se que os pólipos desta espécie não apresentam preferência por nenhum dos substratos em estudo, apresentando ao final de 7 dias uma taxa de fixação superior a 90% independentemente dos substratos usados. Definido o substrato a utilizar nos ensaios, procedeu-se ao estudo das condições ótimas de temperatura e alimentação na reprodução assexuada por gemulação, tendo sido estudadas três temperaturas (20, 24 e 26ºC), combinadas com duas condições de alimento (com e sem alimentação). Verificou-se que a produção de novos pólipos por gemulação obtém-se fornecendo alimento aos pólipos e mantendo-os a 24 ou 26ºC. Do ponto de vista de desenvolvimento de éfiras em pequenas medusas, testaram-se diferentes alimentações ((A) náuplios de artémia (10 Ind/ml), (B) rotíferos (20 Ind/ml), (C) náuplios de artémia (5 Ind/ml) + rotíferos (10 Ind/ml), (D) Microalga (Nannochloropsis sp., 5x105células/ml), (E) Microalga (Nannochloropsis sp., 5x105células/ml) + náuplios de artémia (10 Ind/ml), (F) Microalga (Nannochloropsis sp., 5x105células/ml) + rotíferos (20Ind/ml)), verificando-se que a utilização de Microalgas + Rotíferos, maximiza o crescimento e a sobrevivência, comparativamente com as restantes dietas em estudo. O presente trabalho permitiu aumentar o conhecimento sobre a reprodução assexuada de pólipos e crescimento de éfiras de P. punctata. Ao longo do trabalho foi possível observar que existem condições chave que permitem otimizar o processo de produção desta espécie em aquacultura. Especialmente a importância da alimentação dos pólipos durante o processo de reprodução assexuada por gemulação, bem como da alimentação das éfiras com microalga e rotíferos conjuntamente.

Highly discriminatory single-nucleotide polymorphism interrogation of Escherichia coli by use of allele-specific real-time PCR and eBURST analysis

In total, 782 Escherichia coli strains originating from various host sources have been analyzed in this study by using a highly discriminatory single-nucleotide polymorphism (SNP) approach. A set of eight SNPs, with a discrimination value (Simpson's index of diversity [D]) of 0.96, was determined using the Minimum SNPs software, based on sequences of housekeeping genes from the E. coli multilocus sequence typing (MLST) database. Allele-specific real-time PCR was used to screen 114 E. coli isolates from various fecal sources in Southeast Queensland (SEQ). The combined analysis of both the MLST database and SEQ E. coli isolates using eight high-D SNPs resolved the isolates into 74 SNP profiles. The data obtained suggest that SNP typing is a promising approach for the discrimination of host-specific groups and allows for the identification of human-specific E. coli in environmental samples. However, a more diverse E. coli collection is required to determine animal- and environment-specific E. coli SNP profiles due to the abundance of human E. coli strains (56%) in the MLST database.

Development and validation of novel methods for microbial source tracking based on Escherichia coli as an indicator of water quality

Microbial pollution in water periodically affects human health in Australia, particularly in times of drought and flood. There is an increasing need for the control of waterborn microbial pathogens. Methods, allowing the determination of the origin of faecal contamination in water, are generally referred to as Microbial Source Tracking (MST). Various approaches have been evaluated as indicatorsof microbial pathogens in water samples, including detection of different microorganisms and various host-specific markers. However, until today there have been no universal MST methods that could reliably determine the source (human or animal) of faecal contamination. Therefore, the use of multiple approaches is frequently advised. MST is currently recognised as a research tool, rather than something to be included in routine practices. The main focus of this research was to develop novel and universally applicable methods to meet the demands for MST methods in routine testing of water samples. Escherichia coli was chosen initially as the object organism for our studies as, historically and globally, it is the standard indicator of microbial contamination in water. In this thesis, three approaches are described: single nucleotide polymorphism (SNP) genotyping, clustered regularly interspaced short palindromic repeats (CRISPR) screening using high resolution melt analysis (HRMA) methods and phage detection development based on CRISPR types. The advantage of the combination SNP genotyping and CRISPR genes has been discussed in this study. For the first time, a highly discriminatory single nucleotide polymorphism interrogation of E. coli population was applied to identify the host-specific cluster. Six human and one animal-specific SNP profile were revealed. SNP genotyping was successfully applied in the field investigations of the Coomera watershed, South-East Queensland, Australia. Four human profiles [11], [29], [32] and [45] and animal specific SNP profile [7] were detected in water. Two human-specific profiles [29] and [11] were found to be prevalent in the samples over a time period of years. The rainfall (24 and 72 hours), tide height and time, general land use (rural, suburban), seasons, distance from the river mouth and salinity show a lack of relashionship with the diversity of SNP profiles present in the Coomera watershed (p values > 0.05). Nevertheless, SNP genotyping method is able to identify and distinquish between human- and non-human specific E. coli isolates in water sources within one day. In some samples, only mixed profiles were detected. To further investigate host-specificity in these mixed profiles CRISPR screening protocol was developed, to be used on the set of E. coli, previously analysed for SNP profiles. CRISPR loci, which are the pattern of previous DNA coliphages attacks, were considered to be a promising tool for detecting host-specific markers in E. coli. Spacers in CRISPR loci could also reveal the dynamics of virulence in E. coli as well in other pathogens in water. Despite the fact that host-specificity was not observed in the set of E. coli analysed, CRISPR alleles were shown to be useful in detection of the geographical site of sources. HRMA allows determination of ‘different’ and ‘same’ CRISPR alleles and can be introduced in water monitoring as a cost-effective and rapid method. Overall, we show that the identified human specific SNP profiles [11], [29], [32] and [45] can be useful as marker genotypes globally for identification of human faecal contamination in water. Developed in the current study, the SNP typing approach can be used in water monitoring laboratories as an inexpensive, high-throughput and easy adapted protocol. The unique approach based on E. coli spacers for the search for unknown phage was developed to examine the host-specifity in phage sequences. Preliminary experiments on the recombinant plasmids showed the possibility of using this method for recovering phage sequences. Future studies will determine the host-specificity of DNA phage genotyping as soon as first reliable sequences can be acquired. No doubt, only implication of multiple approaches in MST will allow identification of the character of microbial contamination with higher confidence and readability.

Human-specific E. coli single nucleotide polymorphism (SNP) genotypes detected in a South-East Queensland waterway, Australia

The World Health Organization recommends that the majority of water monitoring laboratories in the world should test for E. coli daily since thermotolerant coliforms and E. coli are key indicators for risk assessment of recreational waters. Recently, we developed a new SNP method for typing E. coli strains, by which human-specific genotypes were identified. Here, we report the presence of these previously described specific SNP profiles in environmental water, sourced from the Coomera River, located on South East Queensland, Australia, over a period of two years. This study tested for the presence of human-specific E. coli to ascertain whether hydrologic and anthropogenic activity plays a key role in the pollution of the investigated watershed or whether the pollution is from other sources. We found six human-specific SNP profiles and one animal-specific SNP profile consistently across sampling sites and times. We have demonstrated that our SNP genotyping method is able to rapidly identify and characterise human- and animal-specific E. coli isolates in water sources.

Refugee Mental Health Interventions

There is great diversity in the type of interventions carried out under the rubric of “refugee mental health.” This is partly due to the holistic ecological and psychosocial approaches that have come to dominate research and humanitarian understandings of refugee mental health. The diverse application of psychosocial principles in refugee mental health is also extended by the many varied locations in which such interventions are carried out. Guidelines have been developed to aid would-be practitioners of mental health care amongst refugee communities. However, challenges remain in demonstrating the effectiveness of the approaches used. The maxim “do no harm” which must guide all interventions in this area has nonetheless been threatened at times by well-meaning, yet misguided actions. Despite these issues, there is much promise that together with refugees themselves, steps can be taken to promote well-being and relieve distress in communities of people displaced by conflict.

The thermal decomposition of hydronium jarosite and ammoniojarosite

The thermal decomposition of hydronium jarosite and ammoniojarosite was studied using thermogravimetric analysis and mass spectrometry, in situ synchrotron X-ray diffraction and infrared emission spectroscopy. There was no evidence for the simultaneous loss of water and sulfur dioxide during the desulfonation stage as has previously been reported for hydronium jarosite. Conversely, all hydrogen atoms are lost during the dehydration and dehydroxylation stage from 270 to 400 °C and no water, hydroxyl groups or hydronium ions persist after 400 °C. The same can be said for ammoniojarosite. The first mass loss step during the decomposition of hydronium jarosite has been assigned to the loss of the hydronium ion via protonation of the surrounding hydroxyl groups to evolve two water molecules. For ammoniojarosite, this step corresponds to the protonation of a hydroxyl group by ammonium, so that ammonia and water are liberated simultaneously. Iron(II) sulfate was identified as a possible intermediate during the decomposition of ammoniojarosite (421–521 °C) due to a redox reaction between iron(III) and the liberated ammonia during decomposition. Iron(II) ions were also confirmed with the 1,10-phenanthroline test. Iron(III) sulfate and other commonly suggested intermediates for hydronium and ammoniojarosite decomposition are not major crystalline phases; if they are formed, then they most likely exist as an amorphous phase or a different low temperature phases than usual.

Genome analysis reveals insights into physiology and longevity of the Brandt’s bat Myotis brandtii

Bats account for one-fifth of mammalian species, are the only mammals with powered flight, and are among the few animals that echolocate. The insect-eating Brandt’s bat (Myotis brandtii) is the longest-lived bat species known to date (lifespan exceeds 40 years) and, at 4–8 g adult body weight, is the most extreme mammal with regard to disparity between body mass and longevity. Here we report sequencing and analysis of the Brandt’s bat genome and transcriptome, which suggest adaptations consistent with echolocation and hibernation, as well as altered metabolism, reproduction and visual function. Unique sequence changes in growth hormone and insulin-like growth factor 1 receptors are also observed. The data suggest that an altered growth hormone/insulin-like growth factor 1 axis, which may be common to other long-lived bat species, together with adaptations such as hibernation and low reproductive rate, contribute to the exceptional lifespan of the Brandt’s bat.