Lectin histochemistry and ultrastructure of rainbow trout Oncorhynchus mykiss kidneys affected by proliferative kidney

Abstract

Eleven biotinylated lectins and the avidin-biotin-peroxidase system (ABC) were used to detect and localize specific carbohydrate residues of the primary (PKX-PCs) and daughter cells (PKKDCs) of the PKX myxosporean, lymphocytes, macrophages and renal tubular epithelial cells of rainbow trout Oncorhynchus mykiss affected by proliferative kidney disease (PKD). Electron microscopy revealed the presence of the typical P m -P C s and PKX-DCs in the renal interstitium, between tubular epithelial cells, and of some intraluminal forms, and showed that the principal inflammatory cells surrounding PKX were lymphocytes and macrophages. The latter were most often elongated and closely adherent to the PKX cell surface. Except Ulex europaeus agglutinin-I, which was always negative, all the lectins exhibited a specific binding pattern on the cells examined. Concanavalia ensiformis (Con-A), Lycopersicon esculentum (LEA), Lens culinaris (LCA), Phytolacca americana (PWM), Ricinus communis (RCA-I), and Triticum vulgaris (WGA) agglutinins stained PKX cells, macrophages, lymphocytes, and tubular epithelial cells. However, PKX-DCs and intralurninal forms were less intensely stained than PKX-PCs with Con-A, LEA, PWM, and RCA-I, and receptors for LCA and WGA were not detected In the renal interstitium, Arachis hypogea agglutinin (PNA) bound strongly to the cytoplasm of macrophages but much less intensely to PKX-PCs and some lymphocytes. Griffonia simplinfolia agglutinin-I (GS-I) stained intensely the cytoplasm of PKX-PCs and the lumina1 surface of tubular epithelial cells whereas a weak reaction was present in the cytoplasm of PKX-DCs and intraluminal forms Dohchos biflorus and Glycine max agglutinins stained PKX-PCs weakly and inconsistently. These observations reveal some differences in glycoconjugate contents between PKXPCs and PKX-DCs and provide preliminary data on their carbohydrate components. The lectin PNA could be an important tool to evaluate the macrophage reaction during PKD and other infections of fishes. Finally, the electlve stainlng of GS-I for PKX cells may be used in quantitative determinations by light microscopy of the degree of PKX ~nfection, and for detection of the myxosporean when in low numbers. In addition, staining of PKX with lectins may allow recognition of forms of the parasites presently not well known, such as the early infective stages.

Tetracapsuloides bryosalmonae and PKD in juvenile wild salmonids in Denmark

“…Lectin histochemical staining of Tetracapsuloides bryosalmonae cells was performed following a previously published protocol (Castagnaro et al 1991) with a few simplifications: paraffin-embedded kidney sections were deparaffinised and gradually transferred to distilled water, and endogenous peroxidise activity was then blocked by incubating kidney sections in 3% H 2 O 2 in phosphate-buffered saline (PBS) for 10 min at room temperature. The slides were rinsed for 5 min in tap water, and kidney sections were covered with 30 µg ml -1 biotinylated Griffonia simplicifolia lectin (BS-I, Sigma L-3759) for 1 h in a moist chamber.…”

Antigenic and biochemical study of PKX, the myxosporean causative agent of proliferative kidney disease of salmonid fish

“…Both rabbit ant~sera and MAbs produced in this work strengthen the panel of a\ '111able PKX-specific biologics that were previously represented only by GS-1 lectin (Castagnaro et al 1991) and MAb B12 (Adams et al 1992). Our present state of knowledge on PKX antigens is limited to that which is known about myxosporeans in general, for which only 2 biologies, a rabbit antiserum against Myxobolus cerebralis (Hamilton & Canning 1988) and a MAb to Ceratornyxa shasta (Bartholomew et al 1989), are currently available.…”

“…For this reason, probes for PKX have been developed recently. Griffonia simplicifolia 1 (GSI) lectln, whlch recognizes methyl U-D-galactopyranoside on PKX glycoconjugates (Castagnaro et al 1991), and a monoclonal antibody (MAb) (Adams et al 1992) have been used to identify the PKD agent in the iorrn of PKX cells in tissues from salmonid hosts in particular rainbow trout Oncorhynchusmykiss, chinook salmon 0. tshawytscha, coho salmon 0 . kisutch, brown trout Salmo trutta and Atlantic salmon S. salar (Marin d e Mateo et al 1993).…”

Rapid diagnosis of Tetracapsuloides bryosalmonae, the causative agent of proliferative kidney disease (PKD) in salmonid fish by a novel DNA amplification method, loop-mediated isothermal amplification (LAMP)

“…Previously, the diagnosis of PKD was dependent on the history of the fish stock, the geographical location, the demonstration of external and internal signs, and histological examination by light microscope for detection of developmental stages of T. bryosalmonae in the kidney, liver and spleen ). Subsequently, to improve the detection of the PKD, several diagnostic tools were developed and applied, including the use of lectins histochemistry (Castagnaro et al 1991); monoclonal antibodies (Adams et al 1992;Saulnier and De Kinkelin 1996), Polymerase chain reaction (Saulnier and de Kinkelin 1997; Kent et al 1998) and in situ hybridization (Morris et al 1999). Although, these nucleic acid amplification methods can detect even low amounts of T. bryosalmonae DNA, there are some obstacles that prevent the wide use of these methods in relatively small-scale clinical laboratories, for example, the need for precision instruments for amplification or elaborate methods for the detection of amplified products.…”

“…The traditional May-Grunwald -Giemsa stained kidney imprints were used to identify the extrasporogonic stages of the parasite within the kidney (Klontz and Chacko 1984). Also, more specific staining methods like lectins were used to localize and identify the parasite (Castagnaro et al 1991). Diagnosis by tissue sections staining is timeconsuming, requiring more than 2 days for completion.…”