Histological approaches for high-quality imaging of zooplanktonic organisms

Melo, Rossana C. N.; ROSA, Priscila Dallé; Noyma, Natália Pessoa; Pereira, Wânia F.; Tavares, Luiz Eduardo Roland; Parreira, Gleydes G.; Chiarini-Garcia, Hélio; Roland, Fábio

doi:10.1016/j.micron.2007.05.002

Cited by 7 publications

(4 citation statements)

References 38 publications

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“…Bacteria were detected on the exoskeleton on most investigated copepods (n = 25), but gut bacteria were found in only three specimens. The occurrence of bacteria in the lumen of the gut may, however, have been underestimated, because copepods embedded in paraffin tend to lose content of the gut lumen [ 48 ]. The presence of numerous bacteria of varying morphology attached the exoskeleton was furthermore demonstrated through SEM ( Fig 1 ), confirming previous findings by other authors [ 19 ].…”

Section: Discussionmentioning

confidence: 99%

Host-Specific and pH-Dependent Microbiomes of Copepods in an Extensive Rearing System

et al. 2015

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Copepods are to an increasing extent cultivated as feed for mariculture fish larvae with variable production success. In the temperate climate zone, this production faces seasonal limitation due to changing abiotic factors, in particular temperature and light. Furthermore, the production of copepods may be influenced by biotic factors of the culture systems, such as competing microorganisms, harmful algae, or other eukaryotes and prokaryotes that may be non-beneficial for the copepods. In this study, the composition of bacteria associated with copepods was investigated in an extensive outdoor copepod production system. Light microscopy and scanning electron microscopy revealed that bacteria were primarily found attached to the exoskeleton of copepods although a few bacteria were also found in the gut as well as internally in skeletal muscle tissue. Through 16S rRNA gene-targeted denaturing gradient gel electrophoresis (DGGE) analysis, a clear difference was found between the microbiomes of the two copepod species, Acartia tonsa and Centropages hamatus, present in the system. This pattern was corroborated through 454/FLX-based 16S rRNA gene amplicon sequencing of copepod microbiomes, which furthermore showed that the abiotic parameters pH and oxygen concentration in rearing tank water were the key factors influencing composition of copepod microbiomes.

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Section: Discussionmentioning

confidence: 99%

Host-Specific and pH-Dependent Microbiomes of Copepods in an Extensive Rearing System

et al. 2015

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“…The ultrastructure of C. raciborskii was analyzed here in detail by TEM. We used a method of pre-inclusion in agar with the aims of maintaining the optimal preservation of cells and of reducing artifacts caused by mechanic damage and the loss of specimens during sample manipulation ( Melo et al, 2007 ; Silva et al, 2014 ). Untreated cells exhibited subcellular details such as an elaborate three-layered envelope, and a cytoplasm containing an intricate membrane system (thylakoids), with associated phycobilisomes and other structures such as polyphosphate granules and carboxysomes ( Figures 3 and 4 ).…”

Section: Discussionmentioning

confidence: 99%

Potential effects of UV radiation on photosynthetic structures of the bloom-forming cyanobacterium Cylindrospermopsis raciborskii CYRF-01

et al. 2015

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Cyanobacteria are aquatic photosynthetic microorganisms. While of enormous ecological importance, they have also been linked to human and animal illnesses around the world as a consequence of toxin production by some species. Cylindrospermopsis raciborskii, a filamentous nitrogen-fixing cyanobacterium, has attracted considerable attention due to its potential toxicity and ecophysiological adaptability. We investigated whether C. raciborskii could be affected by ultraviolet (UV) radiation. Non-axenic cultures of C. raciborskii were exposed to three UV treatments (UVA, UVB, or UVA + UVB) over a 6 h period, during which cell concentration, viability and ultrastructure were analyzed. UVA and UVA + UVB treatments showed significant negative effects on cell concentration (decreases of 56.4 and 64.3%, respectively). This decrease was directly associated with cell death as revealed by a cell viability fluorescent probe. Over 90% of UVA + UVB- and UVA-treated cells died. UVB did not alter cell concentration, but reduced cell viability in almost 50% of organisms. Transmission electron microscopy (TEM) revealed a drastic loss of thylakoids, membranes in which cyanobacteria photosystems are localized, after all treatments. Moreover, other photosynthetic- and metabolic-related structures, such as accessory pigments and polyphosphate granules, were damaged. Quantitative TEM analyses revealed a 95.8% reduction in cell area occupied by thylakoids after UVA treatment, and reduction of 77.6 and 81.3% after UVB and UVA + UVB treatments, respectively. Results demonstrated clear alterations in viability and photosynthetic structures of C. raciborskii induced by various UV radiation fractions. This study facilitates our understanding of the subcellular organization of this cyanobacterium species, identifies specific intracellular targets of UVA and UVB radiation and reinforces the importance of UV radiation as an environmental stressor.

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“…In addition to the use of an appropriate fixative, the use of a resin, such as LRW, instead of paraffin, was critical to avoid tissue damage induced by heating required for the embedding step with paraffin (Melo et al, 2007; O’Malley et al, 2009).…”

Section: Discussionmentioning

confidence: 99%

“…Thus, the mixture of both aldehydes, used in the present study, which is an adaptation of the method described previously by Karnovsky (1965), was essential to preserve the germ cells. In addition to the use of an appropriate fixative, the use of a resin, such as LRW, instead of paraffin, was critical to avoid tissue damage induced by heating required for the embedding step with paraffin (Melo et al, 2007;O'Malley et al, 2009).…”

Section: Discussionmentioning

confidence: 99%

A New Approach for Optimal Morphological Identification and Immunolabeling of Spermatogonial Cells

et al. 2014

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High quality fixation often inactivates epitopes and gentler fixation can fail to preserve biological structure at the required resolution. For studies of male reproduction, immunofluorescence techniques using paraformaldehyde fixation associated with paraffin as an embedding medium gives good epitope preservation, although the cell becomes morphologically compromised. On the other hand, glutaraldehyde associated with a plastic resin has been used with success to recognize and distinguish each spermatogonial cell subtype, but the antigenic sites become inaccessible to antibodies. Here we describe a new method that provides excellent morphological details of testicular cells while preserving the binding capacity of epitopes. Using a combination of glutaraldehyde and paraformaldehyde as a fixative and LR White resin for embedding, we show that it is possible to clearly recognize spermatogonial subtypes (Aund, A-A4, In and B spermatogonia) on 1-μm thick-sections and to label epitopes such as bromodeoxyuridine, a marker used for cellular cycle studies in the testis. The information gained from this procedure can be critical for understanding spermatogonial process of self-renewal and differentiation.

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Histological approaches for high-quality imaging of zooplanktonic organisms

Cited by 7 publications

References 38 publications

Host-Specific and pH-Dependent Microbiomes of Copepods in an Extensive Rearing System

Host-Specific and pH-Dependent Microbiomes of Copepods in an Extensive Rearing System

Potential effects of UV radiation on photosynthetic structures of the bloom-forming cyanobacterium Cylindrospermopsis raciborskii CYRF-01

A New Approach for Optimal Morphological Identification and Immunolabeling of Spermatogonial Cells

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