Microorganisms are of great importance for the large scale elemental cycles and overallfunctioning of most natural ecosystems, and this also includes the ecology and maintenance offreshwater resources. Anthropogenic actions as well as climate change has greatly affectedfreshwaters and it is therefore important to understand how microorganisms react to suchenvironmental changes. I investigated how one such pressure, increased nutrient levels,influenced freshwater microbial communities and their potential to degrade the globallyabundant biopolymer chitin. To assess the effects of changed nutrient levels on functionalsubcommunities within the natural microbiota, I established a collection of mixed culturesoriginating from Lake Erken and two mesocosms from the same lake subjected to either highor low nutrient amendments. I observed that higher nutrient addition greatly increasedbacterial cell numbers in the source community. However, for the emerging mixed culturesgrowing on chitin as a substrate, those originating from the “Low” nutrient amendmentmesocosm treatment featured higher cell growth potential compared to cultures originatingfrom the “High” ones or inoculated with the natural lake water. Moreover, mixed culturesfrom the mesocosms presented higher chitinase extracellular enzymatic activity compared tothe lake cultures. Interestingly, “High” and “Low” mesocosm cultures were quite constrainedin bacterial growth response (low variance for the respective treatment) while the growthpotential in cultures from the lake were much more diverse, indicating a higher degree ofpatchiness and subcommunities with variable ability to profit from chitin as a substrate.Ongoing work will assess how individual microbial lineages react to variable nutrient levelsand how the composition of less diverse but fully functional subcommunities profiting fromchitin will change under such conditions.

The bacterial pathogen Vibrio cholerae is responsible for ongoing cholera outbreaks in Haiti and elsewhere. Association of V. cholerae with the human host is responsible for fatal disease, but the bacteria also reside as natural inhabitants of aquatic environments, commonly attaching as biofilms to chitinous surfaces of copepods and crabs. Prior studies in V. cholerae demonstrated that competence for genetic transformation, a mechanism of horizontal gene transfer (HGT), requires the TfoX regulator protein that is triggered by chitin, and the HapR transcription factor that is made in response to quorum sensing (QS) signals produced by V. cholerae and Vibrios. To define regulatory components connecting extracellular signals to natural competence, I first demonstrated that QS molecules produced by Vibrios within multi-species chitinous biofilms are required for DNA uptake by V. cholerae, confirming the critical role of QS signals in HGT. Second, I identified by transposon-mutagenesis a new positive regulator of competence, CytR (cytidine repressor), only studied prior in E. coli as a regulator of nucleoside scavenging. Specific mutations in V. cholerae CytR impaired expression of competence genes and halted DNA uptake; and the addition of exogenous cytidine had similar affects as predicted in E. coli. V. cholerae and other competent Vibrios encode TfoX, HapR, and CytR, although none of these regulators directly controls genes coding for the DNA uptake apparatus. Thus, these results have uncovered a regulatory network, likely used by many Vibrios, that contains additional factors linking several extracellular chemical molecules (cytidine, chitin, and QS signals) to DNA uptake. My study has begun to define a molecular mechanism by which both environment and genetics contribute to genome evolution for this important marine pathogen.

<p>Equilibrium isotherms and the adsorption kinetics of heavy metals onto chitosan were studied experimentally. Chitosan, a biopolymer produced from crustacean shells, has applications in various areas, particularly in drinking water and wastewater treatment due to its ability to remove metallic ions from solutions. The adsorption capacity of chitosan depends on a number of parameters: deacetylation degree, molecular weight, particle size and crystallinity. The purpose of this work was to study the adsorption of copper, zinc, mercury, and arsenic on chitosan produced from shrimp shells at a laboratory level.</p><p>The experimental work involved the determination of the adsorption isotherms for each metallic ion in a batch system. The resulting isotherms were fitted using the Langmuir model and the parameters of the equation were determined. Kinetic studies of adsorption for different metallic ions at different concentrations and with different particle sizes were performed in batch and column systems. Simplified models such as pseudo-first-order, pseudo-second-order, and intra-particle diffusion equations were used to determine the rate-controlling step. Some preliminary studies were carried out to address the application of chitosan as an adsorbent in the removal of heavy metals or other metallic ions from natural water and wastewater. The regeneration of chitosan was also studied.</p><p>The results showed that the adsorption capacity depends strongly on pH and on the species of metallic ions in the solution. The optimum pH value for the metallic cation adsorption was between 4 and 6, whereas for arsenic adsorption it was about 3. When the pH is not controlled, the adsorption capacity is independent of the initial pH with the solution reaching a final pH of about 7. It was also found that the Langmuir equation described very well the experimental adsorption data for each metallic ion. The adsorption capacity for the metals on chitosan follows the sequence Hg>Cu>Zn>As.</p><p>The study of the adsorption kinetics of these metallic ions shows that the particle size has a significant influence on the metal uptake rate for copper; but that it has only a slight influence on the adsorption rate of zinc and mercury in the range studied. Arsenic adsorption exhibited an interesting behaviour which depends strongly on the pH of the solution; the uptake increased at short adsorption times and then decreased at long times. The analysis of kinetic models showed that the pseudo-second-order adsorption mechanism is predominant, and the overall rate of the metallic ion adsorption process is therefore controlled by adsorption reactions and not by mass transfer for the range of particle sizes examined in this study.</p><p>With regard to the regeneration of chitosan, it was found that sodium hydroxide is a good agent for zinc and arsenic desorption, whereas ammonium sulphate and sodium chloride were the most suitable for copper and mercury desorption, respectively. The ability of chitosan to remove arsenic from natural water, and copper and zinc from mining waste water was verified. The use of these results for designing purposes is a subject for future work.</p>

Septins are a highly conserved family of eukaryotic proteins having significant homology within and among species. In the budding yeast, Saccharomyces cerevisiae, a septin-based hierarchy of proteins is required to localize chitin in the bud neck prior to septum formation. However, this process has not been clarified in a filamentous, conidiogenous fungus capable of yeast growth, such as Wangiella dermatitidis, a polymorphic agent of human phaeohyphomycosis. Prior studies of this melanized mold showed that some chitin synthase mutants (wdchsΔ) have defects in yeast septum formation, suggesting that the septins of W. dermatitidis might functionally associate with some of its chitin synthases (WdChsp). To test this hypothesis, four vegetative septin homologs of S. cerevisiae were cloned from W. dermatitidis and designated WdCDC3, WdCDC10, WdCDC11, and WdCDC12. Of the four, only WdCDC3 functionally complemented completely a strain of S. cerevisiae with a ts mutation in the corresponding gene, although WdCDC12 did so partially. Functional characterizations by mutagenesis of the four W. dermatitidis septin genes revealed that resulting mutants (wdcdc[delta]) each had unique defects in yeast growth and morphology, indicating that each septin carried out a distinct function. Furthermore, when a wdcdc10[delta] mutation was introduced into five different wdchs[delta] strains, weak genetic interactions were detected between WdCDC10 and WdCHS3 and WdCHS4, and a strong interaction between and WdCHS5. Cytological studies showed that WdChs5p was mislocalized in some septin mutants, including wdcdc10[delta]. These results confirmed that in W. dermatitidis septins are important for proper cellular morphogenesis, cytokinesis, and especially septum formation through associations with some chitin synthases. / text

The research pursued relates to the establishment of interrelationship between various factors affecting chitin extraction, and combination of optimum levels of factors required to maximize the yield of chitin extraction. Optimization of chitin extraction was carried out using crab, lobster, and shrimp solid wastes. The study was divided into two stages: (a) optimization of chitin extractability with respect to particle size; (b) optimization of demineralization and deproteinization stages in lobster chitin extraction using Response Surface Methodology. Particle size had a significant effect on crab and lobster chitin extractability; a particle size of 2.0mm gave the highest yield in chitin extraction. The mean yield of chitin from crab and lobster at particle size of 2.0mm were 28.8% and 23.2%, respectively. Shrimp chitin extractability was not affected by particle size with the mean yield being 25.2%. Response Surface Methodology was used to determine simultaneous effects of (a) concentration of extractant; (b) shell:extractant ratio; (c) temperature of extraction; and (d) time of extraction on deproteinization and demineralization stages of chitin extraction and hence yield of chitin. All variables had an effect on demineralization and deproteinization yields. Maximum demineralization was predictable by a multi-factor model consisting of a combination of concentration of HCl, shell:extractant ratio, temperature and time of extraction. (Abstract shortened by UMI.)

Chitin is a polysaccharide found in abundance in the shell of crustaceans. In this study, chitin was first extracted from shrimp waste material previously demineralized, using proteolytic enzymes, i.e., chymotrypsin and papain. The conditions used for the deproteinization were optimized with respect to 3 factors, viz., pH, temperature, and enzyme to waste (E/W) ratio, using response surface methodology (RSM). / In a second part of the study, chitosan was prepared from chitin by partial deacetylation and its capacity to preserve whole and headless fresh shrimps was evaluated. The shrimps (Pandalus borealis) were dipped in chitosan (1 and 2% w:v), and stored on ice. Chemical indicators such as pH, drip loss, total volatile bases (TVB), nucleotide degradation, and sensory indicators of appearance (including melanosis) and odor, as well as total microbial counts were monitored during 20 days. / As a third and final part, the antimicrobial properties of chitosan were evaluated using several microorganisms implicated in food spoilage and/or food poisoning outbreaks, especially those associated with fish and seafood products. Chitosan prepared from crab offal and used in the previous study on shrimp preservation was compared with commercially available chitosan from Sigma Chemical Co. (St-Louis, MO). (Abstract shortened by UMI.)

Components of successful pest management programs must be complementary and not antagonistic. This project examined interactions between natural enemies of the black cutworm, Agrotis ipsilon (Hufnagel), an important turfgrass pest, and host plant resistance by endophytic grass. Agrotis ipsilon nucleopolyhedrovirus (AgipMNPV) was examined as a bio-insecticide for controlling A. ipsilon in turfgrass. Fresh (1-week-old) AgipMNPV residues killed 76−86% of neonates hatching from eggs on golf course tees, however, residual control of implanted larvae lasted no more than a few weeks. Combinations of AgipMNPV with adjuvants, such as optical brightener and lignin, failed to accelerate or extend efficacy of the virus. AgipMNPV seems better suited for targeted control of early instars than for season-long control. Several applications per growing season would likely be needed to maintain high enough titers on turfgrass to effectively control cutworms. The addition of a chitin synthesis inhibiting turfgrass fungicide failed to synergize AgipMNPV infectivity to A. ipsilon. Choice tests revealed the fungicide residues to be a mild feeding deterrent, the likely cause of slightly reduced mortality from virus infection seen in field trials. Combination applications in turfgrass might interfere with larval ingestion of a lethal virus dose, resulting in prolonged feeding in the field. I examined how feeding on perennial ryegrass (Lolium perenne) with or without Neotyphodium lolii, its alkaloid-producing fungal endophyte, affects susceptibility of A. ipsilon to AgipMNPV. Feeding on endophytic grass neither compromises nor synergizes infectivity of AgipMNPV in the cutworm midgut. However, reduced consumption or avoidance of less-palatable endophytic grass could decrease ingestion of virus and rates of subsequent mortality in the field. Host feeding on endophytic grass had differing effects on the tachinid fly, Linnaemya comta, a fast-developing solitary parasitoid, and the encyrtid wasp, Copidosoma bakeri, a slow-developing gregarious parasitoid. L. comta development did not appear to be affected when its host fed on endophytic grass; in contrast, C. bakeri suffered negative fitness effects. These results suggest that parasitoid life strategy and taxonomy play a role in endophyte mediated tritrophic interactions.

The shrimp processing industry has to deal with the ever-growing costs associated with the disposal of their residuals. However, investigation into the possibility of making high-value biopolymers (chitin and chitosan) from this waste shows significant potential for developing a chitin and chitosan industry in Quebec. / The pharmaceutical and medicine industry is the target market for high-grade chitosan. Based on the related data of cellulose derivatives (the closest substitute for chitosan), the Bass model was used to forecast the sales of high-grade chitosan in Quebec. It is estimated that the potential market for chitosan in Quebec is worth 37 million dollars (in 1999 prices) cumulative for the next 20 year period and 59 million for Canada. In the first year of marketing chitosan, sales in Canada (high-grade) are expected to reach $3.2 million from which $1.55 million is expected to be generated in Quebec. (Abstract shortened by UMI.)

Zugl.: Aachen, Techn. Hochsch., Diss., 2008.

Submitted by Johnny Rodrigues (johnnyrodrigues@ufcg.edu.br) on 2018-04-17T12:59:30Z No. of bitstreams: 1 RAFAELLA LIMA DE MENEZES - DISSERTAÇÃO PPG-CEMat 2014..pdf: 7131098 bytes, checksum: 662a27be023a1d5cf9b17d3f230efb5c (MD5) / Made available in DSpace on 2018-04-17T12:59:30Z (GMT). No. of bitstreams: 1 RAFAELLA LIMA DE MENEZES - DISSERTAÇÃO PPG-CEMat 2014..pdf: 7131098 bytes, checksum: 662a27be023a1d5cf9b17d3f230efb5c (MD5) Previous issue date: 2014-08-28 / A quitina é o segundo polissacarídeo natural mais abundante depois da celulose. É biodegradável, não tóxico e encontrado em ambiente marinho como microfibrilas cristalinas ordenadas formando componentes no exoesqueleto de artrópodes e nas paredes celulares de alguns fungos e bactérias. Propriedades como a biocompatibilidade, biodegradabilidade e baixa toxicidade exibida pelas fibras de quitina combinadas com propriedades mecânicas adequadas torna estas, candidatas para aplicações como material de sutura. Este trabalho tem como objetivo a obtenção e caracterização físico-química e estrutural, de fibras de quitina para aplicação como fios de sutura, extraída a partir de exoesqueletos de camarões (espécie: de Litopenaeus vannamei). Esta pesquisa foi dividida em duas metodologias para um melhor entendimento, sendo, portanto composto do processo de extração da quitina seguido pelo processo de obtenção das fibras. As concentrações de quitina utilizadas na formação das fibras foram de 0,5%, 0, 85%, 1%. Dissolvidas no solvente DMAc/LiCl2 (5%) e coaguladas em etanol (30%), acetona (70%), água destilada (100%), CaCl2 (2%). O pó de quitina obtido na primeira etapa desta pesquisa foi caracterizado através das técnicas de difração de raios-X (XRD) e espectroscopia de infravermelho com transformada de Fourier (FTIR). Através do DRX foi possível identificar o tipo de quitina obtida como a alfa quitina, isto confirma a estrutura da quitina presente na casca de camarão, o FTIR revelou a ausência de proteína, confirmando que a quitina obtida pode ser utilizada como um biomaterial. As fibras obtidas na segunda etapa desta pesquisa foram caracterizadas por microscopia eletrônica de varredura SEM/EDS, o qual revelou a influência dos banhos de coagulação no diâmetro das fibras, o aumento do pH no banho de coagulação levou a uma neutralização mais rápida das fibras que quitina, ocorrendo um fenômeno conhecido como shrink nas fibras. Testes de intumescimento revelaram a influência dos banhos de coagulação utilizados, as fibras coaguladas com solventes orgânicos, possuíram um menor grau de intumescimento, quando comparadas com as fibras coaguladas em solventes aquosos. Para os ensaios mecânicos de tração têm-se que as fibras na concentração de 0,5% de quitina devido a uma melhor homogeneidade da solução, possuíam propriedades mecânicas mais adequadas quando comparadas com as outras fibras produzidas. Os resultados de viabilidade celular das fibras apresentaram percentagem superior a 90%, evidenciado a ausência do solvente nas fibras produzidas. Através dos ensaios de biodegradação é possível observar uma perda de massa nas fibras somente a partir do quinto dia. Os resultados obtidos demonstram que o objetivo da pesquisa foi alcançado, de produzir fibras de quitina a partir da casca do camarão, além de indicarem a possível aplicação deste material como um fio de sutura. / Chitin is the second most abundant natural polysaccharide after cellulose It is biodegradable, non-toxic and found in the marine environment as ordered crystalline microfibrils forming components in the arthropods exoskeleton and the cell walls of fungi and bacteria. Properties like biocompatibility, biodegradability, low toxicity exhibited by chitin fibers combined with adequate mechanical properties make them candidates for applications such as suture material. This work aims, obtaining and characterize through a physicochemical and structural characterization of chitin fibers for their use as sutures, extracted from the shrimp exoskeletons (species: Litopenaeus vannamei). This research was divided into two methodologies for a better understanding, is composed by the chitin extraction process followed by the fiber process obtention. The chitin concentration used in the fibers formation were 0.5% 0 85% 1%. Dissolved in DMAc / LiCl2 (5%) solvent and coagulated in ethanol (30%), acetone (70%), distilled water (100%), CaCl2 (2%). The chitin powder obtained in the first step of this study was characterized using the X-ray diffraction (XRD) and Fourier Transform Infrared Spectroscopy (FTIR) techniques. By XRD it was possible to identify the chitin obtained as alpha chitin, this confirms the chitin structure present in the shrimp shell, the FTIR revealed the absence of protein, confirming that the chitin obtained can be used as a biomaterial. The fibers obtained in the second step of this study were characterized by SEM / EDS, which revealed the influence of coagulation baths in the fiber diameter, the increased pH in the coagulation bath led to a more rapid neutralization of the chitin fibers, a phenomenon known as shrink. Swelling tests revealed the influence of the coagulation baths, the fibers coagulated with organic solvent, owned a lower swelling degree compared to the fibers coagulated in aqueous solvents. For the tensile test the fibers with a chitin concentration of 0.5% due to a better solution homogeneity had more adequate mechanical properties, compared to the other fibers produced. The results for cell viability showed a percentage of 90%, evidencing the solvent absence in the fiber produced. Through biodegradation tests is possible to observe a mass loss in the fibers only from the fifth day. The results demonstrated that the objective was achieved, to produce chitin fibers from the shell of shrimps, and indicate a possible application of this material as a suture wire.