Hymenolepis diminuta: a comparison between young developing, and small, destrobilated worms in the rat intestine

Newly in vitro excysted tapeworms of Hymenolepis diminuta (Cestoda, Cyclophyllidea), 1- to 3-day-old worms and destrobilated worms from rat intestines were investigated by means of light microscopy (LM) and scanning electron microscopy (SEM). It was found that the scolex of 1- and 2-day-old worms had shallow suckers with smooth brims, while 3-day-old and older worms, including destrobilated worms, had deep suckers with puckered brims. The posterior end of 1- and 2-day-old worms had a central cone-shaped structure not present in 3-day-old and older, or destrobilated worms. The repairing of the posterior end and the protonephridial system after excystation or destrobilation was much the same. Tissue remnants moved into the centre of the posterior end, resulting in an indentation with a pore to the exterior. The indentation and its pore became connected to the emptying canals of the protonephridial system, i.e. they developed into the excretory bladder and pore respectively.     

Myoelectric response of the small intestine to the orad presence of the tapeworm Hymenolepis diminuta.

During its 24-hr migratory cycle in the small intestine, Hymenolepis diminuta is located in the orad part of the small intestine during the early morning hours and then in the caudad part of the small intestine during the late afternoon and early evening. During the later period, tapeworm-induced alterations of interdigestive myoelectric activity, a correlate of smooth muscle contraction or intestinal motility, are most intense in the ileal region. The hypothesis tested was that the tapeworm-induced changes in intestinal motility are local responses of the intestine responding to the close proximity of the lumenally positioned tapeworm and to the nutritional state of the host. The small intestine was monitored before and for 20 days after infection using electrodes implanted on the serosa of the small intestine. Myoelectric recordings were analyzed for the frequency of the normal patterns of interdigestive myoelectric spiking patterns and the altered myoelectric spiking related to tapeworm infection. During the morning hours, when the tapeworms are situated in the orad small intestine, no changes were observed during the normal myoelectric pattern of the digestive phase in any region of the intestine. When examined after the conversion of the digestive to interdigestive phase of motility, only on day 10 postinfection was the interdigestive phase significantly altered. It was concluded that the presence of the tapeworm in the orad small intestine during the satiety stage of the rat causes no changes in the electric events of the small intestine, with the exception of day 10 postinfection. Because tapeworms in the orad small intestine do not induce the tapeworm-altered myoelectric activity observed in the afternoon and evening with caudally positioned tapeworms, tapeworm-altered motility is not simply a response of the small intestine to the local presence of the tapeworm.     

cGMP secreted from the tapeworm Hymenolepis diminuta is a signal molecule to the host intestine

3',5'-Cyclic guanosine monophosphate (cGMP), a well-known intracellular second messenger, is released to the intestinal lumen by the tapeworm, Hymenolepis diminuta. Enzyme-linked immunosorbent assay analysis of tapeworm conditioned media shows that cGMP is released at a constant rate. Multidrug resistant (MDR) proteins are efflux transporters for cyclic nucleotides. Two MDR inhibitors, niflumic acid and zaprinast, inhibit cGMP secretion by tapeworms and change the cGMP localization within the tapeworm tegument, as assessed by immunochemistry. cGMP, normally present throughout the tapeworm tegumental cytoplasm, is absent from the outer cytoplasmic band upon treatment with inhibitors. Inhibition of cGMP secretion by colchicine indicates that cGMP secretion is cytoskeleton dependent. Binding studies of [3H]cGMP to ileal segments of intestine demonstrate 2 saturable, reversible, and high-affinity binding sites. These studies demonstrate that cGMP is secreted from the cestode via a cytoskeleton-dependent mechanism and MDR efflux transporters. In addition, cGMP reaching the intestinal lumen can bind to the mucosa via receptors for cGMP. These data, combined with earlier observations of cGMP altering intestinal motility and slowing lumenal transit, indicate that tapeworms alter the physiology of the host digestive process via the secretion and binding of extracellular cGMP to lumenal receptors in the host intestine.     

Biosynthesis of polyisoprenoid lipids in the rat tapeworm Hymenolepis diminuta

The possible occurrence of isoprenoid lipids in the tapeworm, Hymenolepis diminuta, was investigated by analytic and biosynthetic methods. Two-dimensional thin layer chromatography (TLC) resolved the worm's non-saponifiable lipids into cholesterol, farnesol, and several unknown compounds, two of which migrated with dolichol standards on TLC and reacted with phthalic anhydride, a probe for alcohols; the major compound also exhibited a mass spectrum very similar to that of a dolichol standard. A third unknown compound separable by TLC, apparently a quinone, was intrinsically red, was decolorized by treatment with sodium dithionite and migrated on TLC in a more polar position than either ubiquinone-50 or vitamin K1. All three compounds, as well as farnesol, were labelled when worms were incubated with [14C]-mevalonolactone, suggesting that they are endogenous isoprenoids.     

Amino acid metabolism in the rat tapeworm, Hymenolepis diminuta

1. The amino acid metabolism of the rat tapeworm, Hymenolepis diminuta was investigated. 2. In addition to the characteristic end products of helminth metabolism, H. diminuta also forms substantial amounts of 14C-alanine during incubations in 14C-glucose. 3. Of 10 amino acids tested, only 14C-labelled asparate and, to a lesser extent alanine, generated substantial amounts of 14CO2 when incubated with H. diminuta. 4. 14C-aspartate was incorporated into both succinate and acetate, major products of the worms mitochondrial metabolism, but the rates were low when compared to the metabolism of exogenous glycogen. 5. These results suggest that amino acid metabolism in H. diminuta is very limited.     

Galactose utilization by the rat tapeworm, Hymenolepis diminuta.

1. Hymenolepis diminuta incorporated label from 14C-galactose into glycogen, but the sugar would not support net glycogen synthesis. Glucose stimulated the incorporation of label from 14C-galactose into glycogen, while glycerol did not. 2. During incubations in galactose, large internal pools of galactose and galactose 1-P accumulated, while the concentration of glucose 6-phosphate remained unchanged. 3. In vitro culture experiments indicated that galactose would not support worm growth. Therefore, while galactose can be metabolized to a limited extent, it cannot substitute for glucose as a nutrient source.     

Identification of ADPR-transferase activity in the rat tapeworm, Hymenolepis diminuta

1. The nuclear fraction of the rat tapeworm Hymenolepis diminuta (Cestoda) contains the enzyme adenosine diphosphoribosyl transferase (ADPR-transferase). 2. The enzyme catalyzes the postsynthetic modification of some nuclear proteins by the covalent attachment of the (ADP-ribose) moiety of NAD to such proteins. 3. The reaction is dependent on DNA which contains strand-breaks, and chain lengths equivalent to (ADP-ribose) is estimated. 4. The formation of polynucleotide products was competitively inhibited by 3-acetamidobezamide, with a Km of 125 microM. 5. The catalytic properties of ADPR-transferase in Hymenolepis diminuta are similar to those in T. brucei.     

Hymenolepis diminuta: lack of pathogenicity in the healthy rat host.

Whether Hymenolepis diminuta (Cestoda: Cyclophyllidea) might affect adversely the growth of its host under normal conditions was studied. Rats were divided into four experimental groups: (1) rats infected with the tapeworm and fed ad libitum, (2) uninfected rats fed ad libitum, (3) and infected rats fed isocalorically with (4) uninfected rats. Growth rates of infected rats did not differ from infected animals. Infected, meal fed rats limited to 15 g synthetic diet/day grew as rapidly as their uninfected counterparts, and infected rats fed ad libitum did not consume more food than the comparable infected group. There were no significant differences in consumption or in excrement produced between groups (1) and (2) and groups (3) and (4). Weights attained by the worms were not affected by mode of host feeding (ad libitum or meal fed), whether expressed as wet or dry weight. Since H. diminuta appears not to affect nutrient utilization or consumption in a healthy, unstressed host, at least on a gross level, it probably should be considered an endocommensal.     

Enzymes of galactose utilization in the rat tapeworm, Hymenolepis diminuta.

1. Crude enzyme preparations from Hymenolepis diminuta contained galactokinase, galactose 1-phosphate uridyl transferase and UDPgalactose 4-epimerase activity, although their specific activities were low. 2. Galactose 1-phosphate non-competitively inhibited galactose phosphorylation. This inhibition, together with the low specific activities of the enzymes in the pathway of galactose utilization, probably accounts for the inadequacy of galactose as a main nutritive carbohydrate for development of the worm.     

Validation of internal controls for gene expression analysis in the intestine of rats infected with Hymenolepis diminuta

The non-invasive parasitic cestode Hymenolepis diminuta induces hypertrophy, hyperplasia and other changes in cell activity in the intestine of rats which are indicated in the expression of mRNA. We have investigated various house-keeping genes (GAPDH, β-actin, 18S and HPRT) and other internal controls (total RNA/unit biomass, total RNA/unit length of intestine) to validate gene expression in the rat intestine after cestode infection and drug-induced neuromodulation. Variation in GAPDH, β-actin, 18S and HPRT expression was observed in rat jejunal tissue according to treatment. Total RNA/unit length of intestine was found to be the most suitable internal control for normalizing target gene mRNA expression in both infected and/or drug-induced rat intestine. This normalization method may be applied to studies of gene expression levels in intestinal tissue where hypertrophy, hyperplasia, rapid growth and cell differentiation generally occur.