Escherichia coli (E. coli) is a common inhabitant of surface waters in the developed and developing worlds. The majority of E. coli cells present in water are not particularly pathogenic to humans; however, there are some present in small proportion that possess virulence genes that allow them to colonize the digestive tract. Pathogenic E. coli causes acute and chronic diarrheal diseases, especially among children in developing countries and in travelers in these locales. The present study, conducted between August 2007 and July 2008, investigated the prevalence and distribution of virulent E. coli strains as either free or attached cells in the final effluents of three wastewater treatment plants located in the Eastern Cape Province of South Africa and its impact on the physico-chemical quality of the receiving water body. The wastewater treatment plants are located in urban (East Bank Reclamation Works, East London), peri-urban (Dimbaza Sewage Treatment Works) and in rural area (Alice Sewage Treatment Works). The effluent quality of the treatment plants were acceptable with respect to pH (6.9-7.8), temperature (13.8-22.0 °C), dissolved oxygen (DO) (4.9-7.8 mg/L), salinity (0.12-0.17 psu), total dissolved solids (TDS) (119-162 mg/ L) and nitrite concentration (0.1-0.4 mg/l). The other xii physicochemical parameters that did not comply with regulated standards include the following: phosphate (0.1-4.0 mg/L); chemical oxygen demand (COD) (5-211 mg/L); electrical conductivity (EC) (237-325 μS/cm) and Turbidity (7.7-62.7 NTU). Results suggest that eutrophication is intensified in the vicinity of the effluent discharge points, where phosphate and nitrate were found in high concentrations. Presumptive E. coli was isolated from the effluent samples by culture-based methods and confirmed using Polymerase Chain Reaction (PCR) techniques. Antibiogram assay was also carried out using standard in vitro methods on Mueller Hinton agar. The viable counts of presumptive E. coli for the effluent samples associated with 180 μm plankton size ranged between 0 – 4.30 × 101 cfu/ml in Dimbaza, 0 – 3.88 × 101 cfu/ml in Alice and 0 – 8.00 × 101 cfu/ml in East London. In the 60 μm plankton size category E. coli densities ranged between 0 and 4.2 × 101 cfu/ml in Dimbaza, 0 and 2.13 × 101 cfu/ml in Alice and 0 and 8.75 × 101 cfu/ml in East London. Whereas in the 20 μm plankton size category presumptive E. coli density varied from 0 to 5.0 × 101 cfu/ml in Dimbaza, 0 to 3.75 × 101 cfu/ml in Alice and 0 to 9.0 × 101 cfu/ml in East London. The free-living presumptive E. coli density ranged between 0 and 3.13 × 101 cfu/ml in Dimbaza, between 0 and 8.0 × 101 cfu/ml in Alice and between 0 and 9.5 × 101 cfu/ml in East London. Molecular analysis successfully amplified target genes (fliCH7, rfbEO157, ial and aap) which are characteristic of pathogenic E. coli strains. The PCR assays using uidA-specific primer confirmed that a genetic region homologous in size to the E. coli uidA structural gene, including the regulatory region, was present in 3 of the E. coli isolates from Alice, 10 from Dimbaza and 8 from East London. Of the 3 E. coli isolates from Alice, 1 (33.3%) was positive for the fliCH7 genes and 3 was positive for rfbEO157 genes. Out of the 10 isolates from Dimbaza, 4 were xiii positive for fliCH7 genes, 6 were positive for the rfbEO157 genes and 1 was positive for the aap genes; and of the 8 isolates from East London, 1 was positive for fliCH7 genes, 2 were for the rfbEO157 genes, 6 were positive for the ial genes. Antimicrobial susceptibility profile revealed that all of the E. coli strains isolated from the effluent water samples were resistant (R) to linezolid, polymyxin B, penicillin G and sulfamethoxazole. The E. coli isolates from Dimbaza (9/10) and East London (8/8) respectively were resistant to erythromycin. All the isolates were found to be susceptible (S) to amikacin, ceftazidime, ciprofloxacin, colistin sulphate, ceftriaxone, cefotaxime, cefuroxime, ertapenem, gatifloxacin, gentamycin, imidazole, kanamycin, meropenem, moxifloxacin, neomycin, netilmicin, norfloxacin and tobramycin. The findings of this study revealed that the Alice wastewater treatment plant was the most efficient as it produced the final effluent with the least pathogenic E. coli followed by the Dimbaza wastewater treatment plant. In addition, the findings showed that the wastewater treatment plant effluents are a veritable source of pathogenic E. coli in the Eastern Cape Province watershed. We suggest that to maximize public health protection, treated wastewater effluent quality should be diligently monitored pursuant to ensuring high quality of final effluents.

Passive sampling, as opposed to the conventional spot or bottle water sampling technique, has shown to be reliable and efficient in monitoring the toxicologically relevant, freely dissolved (e.g. bioavaialable) concentrations of a wide range of organic contaminants in water. At the same time, partitioning controlled delivery (passive dosing; PD) techniques promise to overcome many of the challenges associated with toxicity testing of hydrophobic substances that may bias the interpretation of toxicity data. The present study investigated the feasibility of coupling silicone rubber passive sampling devices (SR-PSDs) with bioassay techniques for both chemical and ecotoxicological assessment of complex mixtures of organic contaminants in the aquatic environment. SR-PSDs were deployed in water at various locations within the Ythan catchment (north east, Scotland, UK), Forth estuary and the Firth of Forth (east coast of central Scotland, UK) for 7 to 9 weeks. Following retrieval, extracts from the SR-PSDs were analysed for dissolved concentrations of a variety of organic contaminants including PAHs and PCBs using GC-MS and GC-ECD respectively and were screened for a wide range of pesticides using GC-MS/MS and LC-MS/MS. The extracts were further evaluated for acute cytotoxicity (i.e. neutral red uptake assay) and EROD induction potential using rainbow trout liver cell line (Oncorhynchus mykiss; RTL-W1) and for phytotoxicity and developmental toxicity potential using algal growth inhibition test (with a marine phytoplankton, Diacronema lutheri) and fish embryo toxicity test (with embryos from zebrafish Danio rerio) respectively. Overall, the individual and total dissolved concentrations of PAHs (ΣPAH40; parent and branched) and PCBs (ΣPCB32; ortho and mono-ortho) measured in water from the Ythan, Forth estuary and Firth of Forth were relatively low compared with other studies using PSDs. A number and level of pesticides, including insecticides, herbicides and fungicides of varying hydrophobicity (log KOWs ~2.25 to ~5.31) were detected in the silicone rubber (SR) extracts from the Ythan catchment, the Forth estuary and the Firth of Forth, suggesting input mainly from agricultural run-off and possibly from direct discharges. No statistically significant (p<0.05) acute cytotoxicity was observed following 48 h exposure of RTL-W1 cells to SR extracts from the Ythan catchment. But, on a sublethal level, for every site, statistically significant EROD activity was observed to some degree following 72 h exposure. In addition, developmental and algal toxicities on embryos of D. rerio and D. lutheri respectively, were measured in all the deployed samples compared with the procedural controls (undeployed samples). Interestingly, extracts of SR-PSDs from the Forth estuary and the Firth of Forth exhibited growth inhibitions on D. lutheri that were similar to those of extracts from the Ythan, even though, fewer numbers of pesticides were detected in the Forth estuary and Firth of Forth than the Ythan. This suggests that pesticides were not solely responsible for the observed effects in the Ythan catchment. To further improve data from toxicity testing of hydrophobic substances, the study identified the use of SR O-rings as a suitable passive dosing format in in vitro toxicity tests and was partially validated through their use in dosing RTL-W1 cells with two individual PAHs and subsequently determining cytotoxicity and EROD-activity.

Lau Wing-Ngar Vivian. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (leaves 135-144). / Abstracts in English and Chinese. / Acknowledgements --- p.i / Abstract --- p.ii / Abbreviations --- p.vi / List of figures --- p.viii / List of tables --- p.xi / Contents --- p.xiii / Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- General introduction --- p.1 / Chapter 1.2 --- Roadside air pollution worldwide and in Hong Kong --- p.2 / Chapter 1.2.1 --- Air quality in Hong Kong --- p.3 / Chapter 1.3 --- Characteristics of particulate matter --- p.9 / Chapter 1.4 --- Composition and sources of particulate matter --- p.11 / Chapter 1.5 --- Toxic effects of particulate matter --- p.12 / Chapter 1.5.1 --- Lung injury --- p.12 / Chapter 1.5.2 --- Cardiovascular injury --- p.15 / Chapter 1.5.3 --- Mutagenesis and carcinogenesis --- p.16 / Chapter 1.6 --- Aims of my study --- p.16 / Chapter 2 --- Toxic Effects of Heavy Metals Ions on Selected Cultured Cell-lines --- p.18 / Chapter 2.1 --- Introduction --- p.18 / Chapter 2.1.1 --- Metals --- p.18 / Chapter 2.1.1.1 --- Cadmium --- p.22 / Chapter 2.1.1.2 --- Chromium --- p.23 / Chapter 2.1.1.3 --- Lead --- p.25 / Chapter 2.1.1.4 --- Zinc --- p.26 / Chapter 2.1.2 --- Metallothioneins --- p.28 / Chapter 2.1.3 --- p53 --- p.31 / Chapter 2.1.4 --- Tumor Necrosis Factor-alpha (TNF-α) --- p.32 / Chapter 2.1.5 --- Aims of this chapter --- p.32 / Chapter 2.2 --- Materials and methods --- p.35 / Chapter 2.2.1 --- Reagents --- p.35 / Chapter 2.2.2 --- Cultured Cell lines --- p.35 / Chapter 2.2.2.1 --- PU5-18 --- p.36 / Chapter 2.2.2.2 --- LL24 --- p.36 / Chapter 2.2.2.3 --- HBE4-E6/E7 --- p.37 / Chapter 2.2.3 --- Cytotoxicity assays --- p.37 / Chapter 2.2.4 --- ELISA assays --- p.40 / Chapter 2.2.4.1 --- ELISA assay ofp53 levels --- p.41 / Chapter 2.2.4.2 --- ELISA assay of TNF-α levels --- p.43 / Chapter 2.2.5 --- MT gene expression studies by Luciferase assay --- p.44 / Chapter 2.2.5.1 --- PCR amplification --- p.44 / Chapter 2.2.5.2 --- 5´ة End modification of PCR amplified DNA --- p.44 / Chapter 2.2.5.3 --- Ligation of DNA fragment to linearized vector --- p.46 / Chapter 2.2.5.4 --- E. coli. transformation by heat shock --- p.46 / Chapter 2.2.5.5 --- PCR sequencing --- p.47 / Chapter 2.2.5.6 --- Transfection of plasmid into HBE4-E6/E7 cells --- p.49 / Chapter 2.2.5.7 --- Data analysis --- p.50 / Chapter 2.3 --- Results and discussion --- p.51 / Chapter 2.3.1 --- Cytotoxicity assays --- p.51 / Chapter 2.3.2 --- Combination effects of metals on cytotoxicity --- p.61 / Chapter 2.3.3 --- p53 --- p.65 / Chapter 2.3.4 --- TNF-α --- p.68 / Chapter 2.3.5 --- MT gene expression studies by Luciferase assay --- p.69 / Chapter 2.4 --- Conclusion --- p.74 / Chapter 3 --- Effects of Polycyclic Aromatic Hydrocarbons (PAHs) on Cultured Cell-lines --- p.75 / Chapter 3.1 --- Introduction --- p.75 / Chapter 3.2 --- Materials and methods --- p.79 / Chapter 3.2.1 --- Reagents --- p.79 / Chapter 3.2.2 --- Cell culture --- p.79 / Chapter 3.2.3 --- AlamarBlue assay --- p.80 / Chapter 3.2.4 --- EROD assay --- p.80 / Chapter 3.3 --- Results and discussion --- p.84 / Chapter 3.4 --- Conclusion --- p.88 / Chapter 4 --- Chemical and Biological Assays on Roadside Dust --- p.89 / Chapter 4.1 --- Introduction --- p.89 / Chapter 4.1.1 --- Composition of particulate matter in Hong Kong --- p.89 / Chapter 4.1.2 --- Metal contents of particulate matter in Hong Kong --- p.91 / Chapter 4.1.3 --- Possible adverse health impacts of particulate matter --- p.94 / Chapter 4.1.3.1 --- In vitro studies using different cell models --- p.94 / Chapter 4.1.3.2 --- In vivo studies using rodents --- p.97 / Chapter 4.1.3.3 --- Epidemiological studies --- p.98 / Chapter 4.1.4 --- Aims of this chapter --- p.100 / Chapter 4.2 --- Materials and methods --- p.101 / Chapter 4.2.1 --- Sampling of roadside dust --- p.101 / Chapter 4.2.2 --- Chemical analysis of roadside dust --- p.104 / Chapter 4.2.2.1 --- Reagents --- p.104 / Chapter 4.2.2.2 --- Total metal contents --- p.105 / Chapter 4.2.2.3 --- Extractable metal contents --- p.105 / Chapter 4.2.3 --- Biological assays --- p.105 / Chapter 4.2.3.1 --- Cell models --- p.106 / Chapter 4.2.3.2 --- Pretreatment of roadside dust --- p.106 / Chapter 4.2.3.3 --- AlamarBlue assay --- p.106 / Chapter 4.2.3.4 --- ELISA assays --- p.108 / Chapter 4.2.3.5 --- Luciferase assay --- p.108 / Chapter 4.3 --- Results and discussion --- p.110 / Chapter 4.3.1 --- Total metal contents --- p.110 / Chapter 4.3.2 --- Extractable metal contents --- p.113 / Chapter 4.3.3 --- AlamarBlue assay --- p.116 / Chapter 4.3.4 --- p53 --- p.122 / Chapter 4.3.5 --- TNF-α --- p.122 / Chapter 4.3.6 --- Luciferase assay --- p.126 / Chapter 4.4 --- Conclusion --- p.129 / Chapter 5 --- General discussion and conclusion --- p.130 / Chapter 6 --- References --- p.135

La maladie d’Alzheimer est actuellement la forme de démence la plus courante. Les causes, les facteurs de risques ainsi que le(s) mécanisme(s) conduisant à l’apparition des symptômes ne sont pas encore clairement connus. Néanmoins, le rôle central du peptide amyloïde (Aβ) dans le développement de la maladie a été démontré au travers de nombreuses recherches et fait actuellement l’unanimité. L’espèce oligomérique d’Aβ est plus précisément pointée doigt comme l’espèce la plus toxique. La formation des oligomers, au cours du processus d’agrégation, conduit à une population hétérogène en termes de taille et morphologies limitant la compréhension actuelle de leur implication dans le processus pathologique ainsi que dans l’initiation de la maladie. <p>Notre étude structurale minutieuse du processus d’agrégation du peptide Aβ démontre la formation d’agrégats dont le degré d’assemblage augmente au cours du temps. Nous avons montré que les agrégats identifiés comme étant des oligomères adoptent une structure en feuillets β antiparallèles. Tandis que l’interconversion de la structure β d’antiparallèle à parallèle conduit à la formation de fibrilles. Sur base de l’interprétation des spectres infrarouges analysés par corrélation à 2 dimensions, nous suggérons que ce changement de conformation est rendu possible grâce aux modifications des liens hydrogènes. En effet, les liens hydrogènes intramoléculaires qui stabilisent la structure antiparallèle des brins β disparaissent en faveur de liens intermoléculaires conduisant à la formation de feuillets β parallèles. De plus, ce changement de conformation requière la rotation des brins β le long de leur axe respectif. <p>Notre travail a pu mettre en avant le rôle central des oligomères dans la pathologie d’une part par leur rôle d’intermédiaires transitoires nécessaires et obligatoires à la formation des fibrilles mais également par la relation étroite qui existe entre leur structure en feuillets β antiparallèles et leur toxicité cellulaire. La modulation et/ou suppression de cette conformation est requise spécifiquement pour réguler leur toxicité et empêcher le processus de mauvais reploiement du peptide conduisant au développement de la maladie. <p>Enfin, nous avons également apporté de nouvelles informations concernant l’implication des membranes biologiques dans le mécanisme de toxicité des oligomères. Nos résultats démontrent que l’interaction du peptide avec un modèle de la membrane biologique ne conduit pas à la déstabilisation de cette dernière. L’hypothèse suggérant la formation de pores et/ou de canaux ioniques comme mécanisme de cytotoxicité est de facto réfutée par notre travail. Néanmoins, nous suggérons que l’interaction du peptide avec les lipides modifie le processus d’agrégation décrit dans la première partie de notre travail. Elle accélère l’étape de nucléation permettant la formation rapide d’oligomères à la surface de la membrane et accentuant ainsi leur probabilité d’interaction avec les protéines membranaires neuronales telles que les récepteurs de neurotransmetteurs./<p>Aggregation of amyloid-β peptides (Aβ1-40 and Aβ1-42) leads to formation of heterogeneous<p>toxic species, oligomers and fibrils, implicated in Alzheimer’s disease. As oligomers were<p>identified as the most cytotoxic entities, our research did focus on their implications in<p>pathology and the Aβ aggregation process which are currently not fully understood.<p>Using ATR-FTIR spectroscopy, we demonstrated that Aβ oligomers adopt an antiparallel β-<p>sheet structure. β-sheet interconversion from antiparallel to parallel seems to be an important<p>step in the Aβ oligomers-to-fibrils transformation. Furthermore, 2-D correlation analysis of<p>infrared spectra recorded during aggregation showed that Aβ isoforms undergo different β-<p>sheet reorganizations explaining their distinct aggregation kinetics. Aβ1-40 misfolding seems<p>to be related to a greater extent of secondary structure changes (increase of β-sheet structure<p>while α-helices and random coil structures content decrease). On the contrary, the same<p>analysis for Aβ1-42 suggests that a possible β-strand ‘rotation’ triggering inter-H bonding<p>formation and stabilizing fibrils may probably explain the antiparallel to parallel β-sheet<p>conversion.<p>We also provided evidence that cytotoxicity is strongly related to the oligomeric antiparallel<p>β-sheet structure of Aβ. The concomitant absence of antiparallel β-sheet structure due to<p>incubation with whey protein-derived peptide hydrolysate strongly suggests that cytotoxicity<p>and β-sheets organization are related.<p>Formation of β-barrel spanning the lipid membrane has been proposed to explain this Aβ<p>structure-toxicity relationship. In the last part of our work, we demonstrated that the<p>interaction of Aβ1-42 with anionic lipid membranes creates and/or stabilizes specific-size<p>oligomers. These oligomers, especially the dodecamer, are known to be the most toxic.<p>Nevertheless, we could not show that these specific oligomers are implicated in membrane<p>destabilization. Further works are needed to separate and study the individual properties of<p>each oligomer. / Doctorat en Sciences / info:eu-repo/semantics/nonPublished

Made available in DSpace on 2014-06-11T19:22:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-04-25Bitstream added on 2014-06-13T20:10:19Z : No. of bitstreams: 1 jacob_cro_me_rcla.pdf: 1789232 bytes, checksum: 4d1aeb62cf1a6763869b8890da8b652a (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Recentemente as abelhas têm sido devidamente valorizadas como importantes polinizadoras de flores silvestres e cultivadas. A densidade populacional de muitos polinizadores tem diminuído devido, principalmente, à intensificação agrícola e ao uso de pesticidas, prejudicando os serviços de polinização. A metodologia clássica para estimar a toxicidade dos produtos químicos para insetos é determinar a dose letal média (DL50) ou a concentração letal média (CL50), podendo então estabelecer doses que sejam mais seguras aos organismos não-alvo ou benéficos. Além dos efeitos de toxicidade aguda, levando a morte das abelhas, doses subletais dos inseticidas podem provocar alterações comportamentais e fisiológicas nos indivíduos, que ao longo do tempo acarretarão em sérios prejuízos na manutenção da colônia. Um dos inseticidas amplamente utilizado é o fipronil, este atua ligando-se aos receptores do ácido gama-aminobutírico (GABA), interrompendo os canais de cloro, resultando na perda de sinalização inibitória neural. Na literatura pode-se encontrar diversos trabalhos que utilizam como modelo principal a abelha Apis mellifera, porém, é importante ressaltar a diversidade existente entre as abelhas nativas no Brasil, os meliponíneos, e sua participação na conservação da biodiversidade, assim como na polinização de áreas de cultivo, o que torna extremamente importante estudos com essa abelha. Com a finalidade de entender como o fipronil interfere morfo e fisiologicamente em abelhas sem ferrão, a região de interesse deste estudo foram os corpos pedunculados, já que estes são centros cerebrais complexos e tidos como local de convergência multisensorial. Para auxiliar no mapeamento metabólico, utilizou-se como marcador a enzima citocromo oxidase e a enzima caspase-3, técnicas utilizadas na observação de atividade neural... / A few decades the bees are considered an important indicator of high environmental sensitivity, and appreciated as important pollinators of wildflowers and cultivated. The population density of many pollinators have declined to harmful levels to pollination services manly due to agricultural intensification and the use of pesticides. The classic methodology of estimating the effects of chemicals for insects is to determine the median lethal dose (LD50) or median lethal concentration (LC50) that can then establish doses that do not harm non-target organisms or beneficial. Besides the effects of acute toxicity, leading to death bees, sublethal doses of insecticides can cause physiological and behavior changes of individuals over time, resulting in serious harm to maintain the colony. One of the widely used insecticides is fipronil, its acts by binding to gamma-aminobutyric acid (GABA) disrupting chloride channels, resulting in loss of inhibitory neural signaling. In the literature one can find several works using as main bee model Apis mellifera, however, it is important to highlight the diversity of Brazilian native bees, the stingless bees, and their participation in biodiversity conversation, as well as in the pollination of cultivated land. In order to understand how fipronil affect morpho and physiologically the stingless bee S. postica, the region of interest in this study were the mushroom bodies, since these are complex brain centers and used as a place of multisensory convergence. This work established the contact LD50 and Ingestion LC50 to the fipronil insecticide for foragers workers stingless bee Scaptotrigona postica in 0.54ng/bee and 0.24ng/μL of the food after 24 hours, respectively, confirming the high toxicity of this phenylpyrazole, in the groups submitted to contact contamination, were identify morphological... (Complete abstract click electronic access below)

Worldwide, water reuse is promoted as an alternative for water scarcity, however, wastewater effluents have been reported as possible contaminants to surface water. The failure of some wastewater treatment processes to completely remove organic matter and some pathogenic microorganisms allows them to initiate infections. This manifests more in communities where surface water is used directly for drinking. To assess water quality, bacteria alone cannot be used as it may be absent in virus-contaminated water. This study was carried out to assess the quality of two wastewater treatment plant effluents from the Eastern Cape Province of South Africa. Physicochemical parameters and microbiological parameters like faecal coliforms, adenovirus, rotavirus, hepatitis A virus, norovirus and enterovirus were evaluated over a projected period of one year. Physicochemical parameters were measured on site using multiparameters, faecal coliforms enumerated using culture-based methods and viruses are detected using both conventional and real-time PCR. Physicochemical parameters like electrical conductivity, turbidity, free chlorine and phosphates were incompliant with the standards set by the Department of Water affairs for effluents to be discharged. Faecal coliform counts were nil for one plant (WWTP-R) where they correlated inversely (P < 0.01) with the high free chlorine. For WWTP-K, faecal coliforms were detected in 27% of samples in the range of 9.9 × 101 to 6.4× 104 CFU/100ml. From the five viruses assessed, three viruses were detected with Rotavirus being the most abundant (0-2034176 genome copies/L) followed by Adenovirus (0–275 genome copies/L) then Hepatitis A virus (0–71 genome copies/L) in the WWTP-K while none of the viruses was detected in WWTP-R. Species B, species C and Adv41 serotypes were detected from the May 2013 and June 2013 samples where almost all parameters were incompliant in the plant. The detection of these viruses in supposedly treated effluents is suggestive of these being the sources of contamination to surface water and therefore renders surface waters unsafe for direct use and to aquatic life. Although real-time PCR is more sensitive and reliable in detection of viruses, use of cell-culture techniques in this study would have been more efficient in confirming the infectivity of the viruses detected, hence the recommendation of these techniques in future projects of this nature.

Cupriavidus metallidurans CH34 is a Gram-negative & / Doctorat en Sciences agronomiques et ingénierie biologique / info:eu-repo/semantics/nonPublished

The ecological position of micro-algae at the base of the aquatic food web makes them critical components of aquatic ecosystems. Their short generation time also makes them useful biological indicators because they respond quickly to changes in environmental condition, enabling timely identification and assessment of water quality changes. The inclusion of micro-algae as indicators in water resource regulation and management in South Africa has started recently, their more extensive use in biomonitoring and ecotoxicology programmes for water resource management would contribute to the South African policy if water resource protection. The standard algal growth inhibition assay with the species Pseudokirchneriella subcapitata is currently used for monitoring toxicity of in-stream and industrial wastewater discharges to freshwater micro-algae. The relevance of the data generated by standard toxicity bioassays has been questioned, since micro-algae in particular are extremely variable in their sensitivity to a range of contaminants and these standard species used may not occur in the local aquatic environment. As a result, international regulatory agencies, have recommended algal growth inhibition tests be changed from a single standard species to tests with a number of species. One recommendation, in addition to the use of standard toxicity tests, is the use of species isolated from the local environment which may be more relevant for assessing site specific impacts. This study investigated the value and application of locally isolated South African freshwater micro-algae in toxicity tests for water resource management and was carried out in three phases. The first phase involved isolating micro-algae from South African aquatic resources. Micro-algae suitable for toxicity testing were identified and selected using as set of criteria. Three (Scenedesmus bicaudatus, Chlorella sorokiniana and Chlorella vulgaris) out of eight successfully isolated species satisfied the prescribed selection criteria and these were selected as potential toxicity test species. The second phase focused on refining and adapting the existing algal toxicity test protocol (the algal growth inhibition assay) for use on the locally isolated algal species. The refinement of the test protocol was achieved by exposing the locally isolated species to reference toxicants in order to assess and compare their growth and sensitivity to the toxicants under the prescribed toxicity test conditions with that of the standard toxicity test species (Pseudokirchneriella subcapitata) and a commercial laboratory species (Chlorella protothecoides). During this phase, one of the three local species (Scenedesmus bicaudatus) was eliminated as a potential toxicity test species due to inconsistent growth. The third phase of the study involved assessing the sensitivity of the two remaining species (C. vulgaris and C. sorokiniana) to a range of toxicants (reference toxicants, salts, effluents and a herbicide) and comparing it to that of the standard toxicity test species P. subcapitata and C. protothecoides. The toxicants were selected based on their relative importance in the South African context, as well as the practicality of using these local micro-algae to routinely determine the impact of these toxicants on local aquatic resources. The growth of the four micro-algae was stimulated by the selected effluents. The standard toxicity test species P. subcapitata was ranked the most sensitive and of the four species to two reference toxicants and two inorganic salts. Chlorella sorokiniana was ranked the most sensitive of the three Chlorella species to two reference toxicants and two inorganic salts. The herbicide stimulated the growth of C. vulgaris while inhibiting the growth of the other species. Pseudokirchneriela subcapitata and C. sorokiniana showed high intra-specific variability in growth, which made it difficult to determine the effective concentrations of the herbicide and therefore compare the sensitivity of the species. This varied response of micro-algal species to toxicants may result in the biodiversity shifts in aquatic ecosystems, and also supports the recommendation of using a battery of different species to support more informed decisions in water resource management.

Water is an essential need that stimulates health and well being. Increase in population size and urbanization negatively affect water resources due to high demands of effluent outputs. Wastewater is an important reservoir for Escherichia coli and can present significant acute toxicity if released into receiving water sources without being adequately treated. E. coli is used as indicator organism for the detection of faecal contamination. These strains have been considered to be one of the primary causes of diarrhoeal infections worldwide. The present study was conducted between September 2012 and June 2013 to assess the prevalence of pathogenic E. coli strains in the final effluents of four wastewater treatment plants in Chris Hani and Buffalo City Municipalities in the Eastern Cape Province of South Africa. Standard membrane filtration technique was used for bacteriological analysis and molecular based technique was used for identification of E. coli pathotypes. The results were recorded in colony forming units/100 ml. Faecal coliforms ranged between 0-9.6×10³ CFU/100 ml for the wwtp-Q and E. coli densities ranged between 0-8.4×10³ CFU/100ml. Faecal coliforms ranged between 4×10²-9.7×10³ CFU/100 ml for wwtp-M and E. coli densities ranged between 1.2×10¹-8.4×10³ CFU/100 ml. The wwtp-E showed to have bacterial counts of faecal coliforms ranging between 4.0×10³-8.2×10³ CFU/100 ml and E. coli densities ranging between 3.5×10¹-7.1×10³ CFU/100 ml. The WWTP-K in this study was only assessed for the presence of E. coli. Faecal coliforms were assessed by the other members of the group. This plant showed to have E. coli densities ranging between 0-7.5×10²CFU/100 ml. A total of 200 presumptive E. coli isolates were subjected to screening by conventional PCR in which (29%) of the wwtp-M isolates were positively identified as E. coli, (16%) of the wwtp-K, (22%) of the wwtp-Q and (34%) of the wwtp-E isolates were positively confirmed as E. coli. A total of 100 randomly selected E. coli isolates were characterised into different pathotypes. (16%) of positive isolates were detected as EPEC and 11% were detected as UPEC strains. There was no detection for the ETEC strains. Antibiotic susceptibility patterns of E. coli strains showed high levels of resistance to Penicillin G, Erythromycin, Tetracycline and Sulfamethoxazole. High levels of Susceptibility were observed in antibiotics such as Chloramphenicol, Amoxicillin and Tetracycline. The results of this study reveal that the plants were above the recommended Standard limit of zero CFU/100 ml for effluents meant to be discharge into receiving water sources. This study reveals inadequacy of the plants studied to produce effluents of acceptable quality.

Cancer is the given name for a group of more than 100 diseases account for 13% of deaths worldwide and 15.1% in Brazil, with estimates of increased mortality. These data show the deficiency of more effective anticancer agents, leading to increased life expectancy or cure. In this context, nature is an alternative to the problem, for harboring a huge biodiversity and Brazil stands out for having the highest diversity of plant species in the world, but little explored as their biological characteristics. The genus Mentha has its very widespread species in the country, with many evidenced therapeutic activities, among them the potential anticancer action of essential oil (EO) demonstrated in two species (Mentha piperita e Mentha spicata). However, this study aimed to evaluate the antitumor activity of the essential oil of Mentha x villosa Hudson (OEMV) in vitro and in vivo, and its toxicity in vivo. For this, the EO was extracted from the leaves of Mentha x villosa, 12 of its constituents have been identified and their main constituent, the monoterpene rotundifolone was isolated. The in vitro cytotoxicity of OEMV rotundifolone and 3 were evaluated against tumor cell lines: ovarian adenocarcinoma (OVACAR-8), colon carcinoma (HCT-116) and glioblastoma (SF295) by the MTT assay. The OEMV showed cytotoxic activity in all cell lines tested, with IC50 ranging from 0.57 to 1.02 ìg/mL. However, rotundifolone did not show cytotoxicity at the tested concentrations, suggesting that the activity of the oil is not mediated by its major constituent. Considering this result, we evaluated the antitumor activity of OEMV in vivo using mice transplanted with sarcoma 180. In this study, it was showed antitumor activity in OEMV treatments performed by intraperitoneal (32.02 and 42.81% with 50 and 100 mg/kg/day, respectively) and orally (34.27 and 43.22% with 100 and 200 mg/kg/day, respectively), probably due to synergy among its constituents. No change was observed in the toxicological parameters evaluated in animals treated with OEMV: variation in body mass, organ weights (liver, spleen and kidney), blood biochemical parameters (AST, ALT, urea and creatinine) and ulcerative lesion index (this later only in the groups treated orally). Since the results demonstrated that OEMV has antitumor activity with low toxicity, it was proposed to evaluate the benefit of the association between OEMV (50 and 100 mg/kg/day) and 5-fluorouracil (5-FU, 10 mg/kg/day) reference known antineoplasic and toxicity. The association promoted tumor growth inhibition of 50.3% and 65.2% at doses of 50 and 100 mg/kg/day, respectively. The highest dose of OEMV in combination showed the same statistical difference that the level of inhibition of 5-FU at the highest dose (25 mg/kg/day). The toxicological parameters, variation of body mass and biochemical parameters (AST and ALT) showed alterations similar as the positive control. The organ weight alterations did not occur, a result that demonstrates an evident benefit of the combination compared to the group treated with 5-FU alone. The last toxicological endpoints evaluated were hematological parameters where both groups showed changes associated with differential count (lymphocytosis and neutropenia) and reduction in total leukocytes; positive factor in this result is that the reduction was not as intense as the one displayed by the control 5-FU furthermore possibly resulting in a decreased susceptibility to infection. The association promoted tumor growth inhibition 50.3% and 65.2% at doses of 50 and 100 mg/kg/day, respectively. The highest dose of OEMV in combination showed the same statistical difference that the level of inhibition of 5-FU dose (25 mg/kg/day) the toxicological parameters, variation of body mass and biochemical analysis (AST and ALT) showed alterations similar to the positive control. / O cancer e o nome dado a um conjunto de mais de 100 doencas, responsaveis por 13% das mortes no mundo e 15,1% no Brasil, com estimativas de mortalidade crescente. Esses dados refletem a carencia por agentes antineoplasicos mais efetivos, que levem ao aumento da expectativa de vida ou a cura. Nesse contexto, a natureza e uma alternativa para o problema, por abrigar uma gigantesca biodiversidade e o Brasil se destaca, por possuir a maior diversidade de especies de plantas do mundo, mas pouco exploradas quanto suas caracteristicas biologicas. O genero Mentha tem suas especies bem difundidas no pais, com diversas atividades terapeuticas comprovadas, dentre elas o potencial acao anticancer do oleo essencial (OE) comprovadas em duas especies (Mentha piperita e Mentha spicata). Portanto, o presente estudo teve como objetivo avaliar a atividade antitumoral do oleo essencial da Mentha x villosa Hudson (OEMV) in vitro e in vivo, e sua toxicidade in vivo. Para isso, o OE foi extraido a partir das folhas da Mentha x villosa, onde 12 dos seus constituintes foram identificados e isolado seu constituinte majoritario, a rotundifolona . A citotoxicidade in vitro do OEMV e da rotundifolona foi avaliada frente a 3 linhagens de celulas tumorais: adenocarcinoma ovariano (OVACAR-8), carcinoma de colon (HCT-116) e glioblastoma (SF295) atraves do ensaio do MTT. O OEMV apresentou atividade citotoxica em todas as linhagens de celulas testadas, com CI50 variando entre 0,57 e 1,02 Êg/mL. Porem, a rotundifolona nao demonstrou atividade citotoxica nas concentracoes testadas, sugerindo que a atividade do oleo nao e mediada por seu constituinte majoritario. Diante desse resultado, foi avaliada a atividade antitumoral do OEMV in vivo utilizando camundongos transplantados com sarcoma 180. Neste ensaio, o OEMV apresentou atividade antitumoral nos tratamentos realizados pela via intraperitoneal (32,02 e 42,81% com 50 e 100 mg/kg/dia, respectivamente) e pela via oral (34,27 e 43,22% com 100 e 200 mg/kg/dia, respectivamente), por provavel sinergismo entre seus constituintes. Nao foi observada nenhuma alteracao nos parametros toxicologicos avaliados em animais tratados com o OEMV: variacao de massa corporea, massa dos orgaos (figado, baco e rins), analises bioquimicas (AST, ALT, ureia e creatinina) e indice de lesao ulcerativa (este ultimo apenas nos grupos tratados por via oral). Como os resultados demonstraram que o OEMV possui atividade antitumoral, com baixa toxicidade, foi proposto avaliar o beneficio da associacao, via i.p, entre o OEMV (50 e 100 mg/kg/dia) e o 5-Fluorouracil em menor dose (5-FU, 10 mg/kg/dia), antineoplasico de referencia e conhecida toxicidade. A associacao promoveu a inibicao do crescimento tumoral de 50,3% e 65,2% nas doses de 50 e 100 mg/kg/dia, respectivamente. A maior dose do OEMV em associacao demonstrou a mesma diferenca estatistica que o indice de inibicao do 5-FU na maior dose (25 mg/kg/dia). Os parametros toxicologicos, variacao de massa corporea e analise bioquimica (AST e ALT) apresentaram alteracoes semelhantes as do controle positivo. Na massa dos orgaos nao ocorreu alteracao, resultado que demonstra um evidente beneficio da associacao quando comparada ao grupo tratado com 5-FU isolado. O ultimo parametro toxicologico avaliado foram as analises hematologicas onde ambos os grupos associados demonstraram alteracoes na contagem diferencial e reducao na contagem de leucocitos totais; o fator positivo nesse resultado e que a reducao nao foi tao intensa quanto a apresentada pelo controle positivo 5-FU, provocando possivelmente, uma menor susceptibilidade a infeccoes. Diante dos resultados, podemos afirmar que o OEMV tem atividade antitumoral in vitro e in vivo com baixa toxicidade, e que a associacao entre 5-FU e o OEMV potencializa a atividade antitumoral do antineoplasico, diminuindo alguns dos seus efeitos toxicos.