Usha Kumari

Comparative analysis of innate immune parameters of the skin mucous secretions from certain freshwater teleosts, inhabiting different ecological niches

The innate immune system of fish is considered first line of defense against a broad spectrum of pathogens. Being a component of innate immunity and lying at the interface between fish and the aqueous environment, skin mucus plays a frontier role in protecting fish from infections. In the present study, skin mucus of Cirrhinus mrigala, Labeo rohita, Catla catla, Rita rita and Channa punctata, inhabiting different ecological niches, was analyzed to characterize potential innate immune factors such as lysozyme, proteases, phosphatases, esterase and sialic acid. The enzyme activities were high in bottom dweller species, C. punctata and C. mrigala, and low in clean water inhabiting species, L. rohita and C. catla. An inverse relationship was observed between the level of enzyme activity and the sialic acid content in these fish species. In R. rita, however, the levels of all factors were found to be low. Zymographic analysis with labeled Micrococcus lysodeikticus revealed three isoforms of lysozyme in C. punctata and two in each species, C. mrigala, L. rohita and C. catla. In R. rita, lysozyme could not be detected. Gelatin zymography revealed that serine and metalloproteases were the major mucus proteases in all fish species investigated. In addition, trypsin-like protease and Ca++-specific serine proteases were observed in skin mucus. Increased knowledge of these parameters could be useful in understanding the role of skin mucus in the innate immune system of fish species inhabiting different ecological niches.

Morphological specializations of the buccal cavity in relation to the food and feeding habit of a carp Cirrhinus mrigala: A scanning electron microscopic investigation

The buccal cavity of an herbivorous fish, Cirrhinus mrigala, was investigated by scanning electron microscopy to determine its surface ultrastructure. The buccal cavity shows significant adaptive modifications in relation to food and feeding ecology of the fish. The buccal cavity of the fish is of modest size and limited capacity, which is considered an adaptation with respect to the small‐sized food items primarily consumed by the fish that could be accommodated in a small space. Modification of surface epithelial cells, on the upper jaw, into characteristic structures—the unculi—is considered an adaptation to browse or scrap, to grasp food materials, e.g., algal felts, and to protect the epithelial surface against abrasions, likely to occur during their characteristic feeding behavior. Differentiation of the highly specialized lamellar organ on the anterior region of the palate could be an adaptation playing a significant role in the selection, retention, and sorting out of palatable food particles from the unpalatable items ingested by the fish. The filamentous epithelial projections and the lingulate epithelial projections on the palatal organ in the posterior region of the palate are considered to serve a critical function in final selection, handling, maneuvering, and propelling the food particles toward the esophagus. The abundance of different categories of taste buds in the buccal cavity suggests that gustation is well developed and the fish is highly responsive in the evaluation and the selection of the preferred palatable food items. The secretions of mucous cells in the buccal cavity are associated with multiple functions—particle entrapment, lubrication of the buccal epithelium and food particles to assist smooth passage of food, and to protect the epithelium from possible abrasion. These morphological characteristics ensure efficient working of the buccal cavity in the assessment of the quality and palatability of ingested food, their retention and transport toward the esophagus. Such an adaptation may be essential in conducting the function most basic to the survival of the individuals and species—feeding. J. Morphol. 2009.

Histochemical analysis of glycoproteins in the secretory cells in the gill epithelium of a catfish, Rita rita (Siluriformes, Bagridae).

Glycoproteins (GPs) were visualised histochemically in the secretory cells - the mucous goblet cells (the type A and the type B), the serous goblet cells, the club cells and the epithelial cells in the gill epithelium of Rita rita. The type A mucous goblet cells, the type B mucous goblet cells and the epithelial cells elaborate GPs with oxidizable vicinal diols and GPs with sialic acid residue without O-acyl substitution. In addition, GPs with O-sulphate esters are elaborated by the type A and GPs with O-acyl sugars by the type B mucous goblet cells. GPs are absent in the serous goblet cells and are with oxidizable vicinal diols in low moieties in the club cells. The analysis of the results elucidates interesting differences in the composition and concentration of GPs in the mucus elaborated by the epithelium of the gill arches and the gill rakers; and the gill filaments and the secondary lamellae indicating the potential importance of the glycoproteins at these locations. GPs elaborated on the surfaces of the gill arches and the gill rakers could be associated to assist in feeding activities and on the surfaces of the gill filaments and the secondary lamellae in the respiratory activity.

Surface ultrastructure of gill arches and gill rakers in relation to feeding of an Indian major carp, Cirrhinus mrigala

The surface ultrastructure of the gill arches and the gill rakers of an herbivorous fish, Cirrhinus mrigala was investigated by scanning electron microscopy. These structures show significant adaptive modifications associated with the food and feeding ecology of the fish. Closely lying short gill rakers and narrow inter-raker channels on the gill arches are associated to filter and retain food particles. Prominent epithelial protuberances on the gill rakers and the gill arches enable the taste buds, located at their summit, to project well above the surface of the epithelium. This could increase the efficiency of the taste buds in selective sorting of palatable food. Surface specializations of the postlingual organ are recognized adaptive modifications for selecting, trapping or holding food particles. Prominent molariform teeth born on the lower pharyngeal jaw, and the chewing pad opposite it, are associated to work together as an efficient pharyngeal mill. Mucous goblet cells are considered to elaborate mucus secretions to trap, glue and lubricate food particles for their smooth transport for swallowing.

The First Evidence of Cholinesterases in Skin Mucus of Carps and its Applicability as Biomarker of Organophosphate Exposure

The presence of cholinesterase (ChE) activity in skin mucus of three carps, Cirrhinus mrigala, Labeo rohita, and Catla catla and its applicability as biomarker of the organophosphorus insecticide exposure were investigated. Biochemical characterization, using specific substrates and inhibitors, indicated that measured esterase activity in skin mucus was mainly owing to ChEs. Significant difference in the proportion of acetylcholinesterase and butyrylcholinesterase activities was observed in skin mucus of three carps. Enzyme kinetic analysis, using the substrate acetylthiocholine iodide revealed significantly high Vmax value in C. catla compared to that in L. rohita and C. mrigala. In contrast, Vmax value using the substrate butyrylthiocholine iodide was significantly high in C. mrigala than in L. rohita and C. catla. In vitro treatment of skin mucus of three carps, with the organophosphorus insecticide Nuvan®, showed strong inhibition of ChE activities. In vivo experiments conducted using C. mrigala and exposing the fish to the sublethal test concentrations (5 and 15 mg/L) of the insecticide also revealed significant inhibition of ChE activity in mucus. In C. mrigala, exposed to the sublethal test concentrations of the insecticide for 4 days and then kept for recovery for 16 days, mucus ChE activity recovered to the control level. Thus, ChE activity in skin mucus could be considered a good biomarker of the organophosphorus insecticide exposure to fish and a useful tool in monitoring environmental toxicity.

Healing of cutaneous wounds in a freshwater teleost, Labeo rohita: Scanning electron microscopical investigation

In this study, healing of cutaneous wounds in Labeo rohita using scanning electron microscope is reported. Wound area could be divided into three regions. Immediately after infliction of wound, edges retract exposing underlying tissues in wound gap (Region I). Simultaneously, at region close to wound edge (Region II), mucous goblet cell openings are observed with copious mucous secretions. Within 1 h, Region I gets covered by mucous secretions, and epidermis at edges starts migrating. Opposing fronts gradually advance and by 4–6 h come in contact to epithelialize wound gap. Zone of contact of fronts is demarcated by epidermal ridge, which is relatively prominent at 8 h. It gradually diminishes and is not distinguished at 24 h and afterward. At 1–4 h, microridges on epithelial cell surfaces appear irregularly arranged, widely spaced, short with abrupt ends at Region I; relatively extensive at Region II; and similar to those in controls at region surrounding Region II (Region III). At 12 h and afterward, microridges appear similar to those in controls at Regions I and II. At 1–2 h, isolated swollen epithelial cells, often in process of detachment and exfoliation at surface, are observed at Regions I and II. Such cells are infrequent at 8 h and afterward. Region I covered by migrated epidermis appears trough like at 4 h to 2 days, level of which gradually rises and at Day 4, surface of epidermis appears at a level similar to that at Regions II and III. Changes have been associated with the imbalance of osmotic homeostasis due to disruption of barrier between internal and external environment of skin. Microsc. Res. Tech. 2012.

Histochemical analysis of glycoproteins in the secretory cells in the epidermis of the head skin of Indian Major Carp, Labeo rohita

A series of histochemical procedures were employed to localise and characterise glycoprotein (GP) classes produced by the epithelial cells, the type A and the type B mucous goblet cells (MGCs) and the club cells in the epidermis of Labeo rohita. The epithelial cells secreted GPs with oxidizable vicinal diols and GPs with sialic acid residues without O-acyl substitution in low concentrations. The type A MGCs and the type B MGCs, in contrast, produced these GPs in high concentrations. Further, these MGCs produced GPs with O-sulphate esters as well. GPs with O-sulphate esters were produced in high concentration by the type A MGCs and in low concentration by the type B MGCs. The club cells produced GPs with oxidizable vicinal diols in trace amounts. Production of more than one type of GPs suggested a basis for functional discrimination in their role in the mucous secretions at the skin surface. This is considered an adaptation to environment inhabited by the fish and is discussed in relation to their role in lubrication, protection and inhibition of the invasion and proliferation of pathogenic micro-organisms.

Histochemical analysis of glycoproteins in the gill epithelium of an Indian major carp, Cirrhinus mrigala

Glycoproteins were analyzed by a range of histochemical methods in the epithelium of gills of Cirrhinus mrigala, a valuable food fish of great economic importance cultured extensively in India. The gills consist of gill arches, gill rakers, gill filaments and secondary lamellae. Major components of the epithelium of gill arches and gill rakers are epithelial cells, mucous goblet cells, rodlet cells, lymphocytes, eosinophilic granular cells and taste buds. In contrast, in the gill filament epithelium, rodlet cells and taste buds, and in secondary lamellae epithelium, rodlet cells, lymphocytes, eosinophilic granular cells and taste buds are not discernible. The epithelial cells, the mucous goblet cells and the eosinophilic granular cells elaborate glycoproteins with oxidizable vicinal diols and glycoproteins with sialic acid residues without O-acyl substitution. In addition, glycoproteins with O-sulphate esters are secreted by the mucous goblet cells. The rodlet cells elaborate glycoproteins with oxidizable vicinal diols. Different types of glycoproteins elaborated on the epithelial surface of gills are discussed in relation to physiological significance of glycoprotein classes with special reference to their roles in lubrication, protection and inhibition of invasion and proliferation of pathogenic micro-organisms.

Response of the chromatophores in relation to the healing of skin wounds in an Indian Major Carp, Labeo rohita (Hamilton).

Chromatophores show significant changes during healing of skin wounds in Labeo rohita (Common Name--Rohu). Wound area can be divided into regions I, II and III. After infliction of wound, skin colour becomes significantly dark by 2h that is gradually restored by 2d. In regions II and III at 5 min, epidermal melanophores appear with beaded dendrites. In these regions at 2h and in region I at 6h, epidermal melanophores appear small, rounded or irregular shaped having dendritic processes with aggregated melanosomes. Subsequently, melanophores appear having elongated dendrites with dispersed or aggregated melanosomes. At 24h, clusters of pigmented bodies appear in regions I and II. These bodies increase up to 2d, and then diminish gradually and disappear by 8d. Changes in dermal melanophores in region II at 5 min indicate the onset of degeneration. Degenerating melanophores increase up to 12h, then gradually decline, and disappear by 4d. Simultaneously, stellate melanophore reappear, gradually increase and appear like control by 8d. Dermal melanophores in region III at different intervals appear stellate. In region I stellate dermal melanophores appear at 4d. Stellate melanophores in all regions show different distribution of dispersed or aggregated melanosomes. With the appearance of dermal melanophores, highly refractive, crystalline structures, possibly the refractive platelets of the iridophores, are visualized around them. At subsequent intervals, these are frequently observed. This study provides interesting insights in injury induced changes in chromatophores in fish. The findings could be considered useful in perception of intriguing features in the development of pigment research in future.

Surface ultrastructure of the gill filaments and the secondary lamellae of the catfish,Rita rita, and the carp, Cirrhinus mrigala

Surface ultrastructures of gill filaments and secondary lamellae of Rita rita and Cirrhinus mrigala, inhabiting different ecological habitat, were investigated to unravel adaptive modifications. R. rita is a sluggish, bottom dwelling carnivorous catfish, which inhabits regions of river with accumulations of dirty water. It retains its viability for long time if taken out of water. C. mrigala is an active bottom dwelling Indian major carp, which lives in relatively clean water and dies shortly after taken out of water. In R. rita, gill septa between gill filaments are reduced. Microridges on epithelial cells covering gill filaments are often continuous and arranged concentrically. Secondary lamellae are extensive. The epithelium appears corrugated, show irregular elevations and shallow depressions, and microridges on epithelial cells appear fragmented. In C. mrigala, in contrast, the gill septa are extensive. Microridges on epithelial cells covering gill filaments are fragmented. Secondary lamellae are less extensive. The epithelium appears smooth and microridges on epithelial cells are relatively inconspicuous. These differences have been considered adaptive modification in relation to habit and ecological niches inhabited by two fish species. Presence of mucous goblet cells on gill filaments is discussed in relation to their functions including precipitation of the sediments and preventing clogging of gill filaments. Infrequent mucous goblet cells in the epithelium of secondary lamellae in two fish species are considered an adaptation, minimizing thickness of the epithelium to reduce barrier between blood and water for favoring gasses exchange with increased efficiency. Microsc. Res. Tech., 2011.