Antigenic differences betweenTrichinella spiralis andT. pseudospiralis detected by monoclonal antibodies

Antigenic differences betweenTrichinella spiralis andT. pseudospiralis were established using two monoclonal antibodies (mAbs) that show different specificities to muscle larvae of the two variants. Enzymelinked immunosorbent assay (ELISA) revealed that mAb 3G6 reacts positively againstT. spiralis, T. nelsoni, T. nativa andT. pseudospiralis, whereas mAb 3E10 does not react withT. pseudospiralis under the same experimental conditions. These antigenic differences were confirmed after preabsorption of the antibodies with serial dilutions of extracts ofT. spiralis orT. pseudospiralis muscle larvae. The indirect immunofluorescence technique showed that the antigen corresponding to mAb 3G6 is located in the stichosomes and the cuticle surface of bothT. spiralis andT. pseudospiralis. In contrast, mAb 3E10 positively stained cryostat sections ofT. spiralis, forming a dense reaction product on the surface of the whole larvae and the surrounding capsule. This antibody can be quite useful as a specific probe for distinguishingT. spiralis fromT. pseudospiralis in taxonomic studies. Using an avidin-biotin system, we could prove that mAb 3G6 recognizes an excretory/secretory-type antigen.     

Protective immunity againstAngiostrongylus cantonensis in rats following sensitizing infections

Three strains (ACI, August and Wistar) of rats previously sensitized by oral infection with intact third-stage larvae ofAngiostrongylus cantonensis developed significant protective immunity to challenge infections 6 weeks later. The degree of the immunity was highest in the August strain of rats, followed by Wistar and ACI rats. Protective immunity appears to affect both third-stage larvae and fourth- and/or fifth-stage worms. ACI rats showed poor antibody responses, especially in the IgE fraction. When 24-day-old young adult worms were transferred from the brain of donor rats into the peritoneal cavity of sensitized rodents, peritoneal eosinophils predominantly adhered to the worm surfaces in vivo.     

Effects of milbemycin D on adult Angiostrongylus cantonensis in rats

Effects of milbemycin D against adult Angiostrongylus cantonensis in rats were examined. The first-stage larval counts in rat faeces (larvae per gram of faeces per female worm recovered, LPG/female) were most conspicuously reduced in the group treated with nine consecutive weekly doses of 5.0 mg/kg. The effect was more marked in the group treated with five or ten successive daily doses of 5.0 mg/kg than the group treated with a single dose of 25.0 or 5.0 mg/kg. Host lung-body weight ratio and number of recovered worms were reduced significantly only in the group treated with five or ten successive daily doses of 5.0 mg/kg. These results suggest that the action of milbemycin D on the reproductive system of the worms might be differentiated from its killing action. The in vitro motility of females recovered from rats medicated with nine consecutive weekly doses of 5.0 mg/kg was inhibited, and almost all females and males were semitransparent and colourless. Results obtained from sectioned worms showed little content in their digestive tracts and uteri. In addition, there were few eggs and first-stage larvae in the lung tissues of treated rats. These suggest that milbemycin D affects the reproductive functions of the worms through an indirect mode of action including paralysis and inhibition in food intake and energy and/or synthetic metabolism.Studies of chemotherapy of parasitic helminths XXIX     

Angiostrongylus cantonensis: An optimized cultivation of this parasitic nematode under laboratory conditions

Parasitology Research - Angiostrongylus cantonensis (A. cantonensis), a parasitic nematode, is the important neurotropic pathogen which causes human angiostrongyliasis. It has a complex life-cycle...     

Angiostrongylus cantonensis: phospholipase in nonsensitized and sensitized rats after challenge.

Rats given an initial infection with Angiostrongylus cantonensis had moderately elevated phospholipase B activity in the lungs at 8 and 15 days after challenge, and greatly elevated levels were evident at 35, 43, and 49 days. In the brain, the values were elevated at 15 through 35 days. These periods of increased activity in the lungs and brain coincided with the migration patterns of the third stage larvae and the adult worms in this host. The elevated enzyme levels also were were correlated with increased numbers of eosinophils in the bone marrow at 8 and 15 days and again at 36, 43, and 49 days after infection. Similarly infected rats exhibited leukocytosis at 1 through 10 weeks of observation after challenge, and striking eosinophilia at 1, 7, 8, and 9 weeks. Rats reinfected after removal of the worms of the initial infection by thiabendazole treatment showed an anamnestic response characterized by (i) elevated enzyme values in both the lungs and brain at 1 day after reinfection and (ii) eosinophilia in the bone marrow by day 4. These accelerated responses were accompanied by a significant reduction in the worm burden of the rats. The results, which support our hypothesis that inflammation, elevated phospholipase B activity, and reduction in worm burden are causally related, are discussed in light of similar findings reported earlier from our studies with Trichinella spiralis and Nippostrongylus brasiliensis.     

广州管圆线虫致大白鼠心脏病变的形态学观察

目的：探讨广州管圆线虫所致大白鼠心脏的病理变化。方法：大白鼠感染管圆线虫后,用光镜与电镜观察其心脏的形态改变。结果：感染30天后的大白鼠右心房室及肺动脉可见成虫寄生,心肌纤维变性,嗜酸粒细胞浸润,内膜细胞脱落,外膜有纤维渗出。电镜下肌膜高度水肿,肌丝溶解,线粒体空泡变等。结论：广州管圆线虫所致大白鼠的心脏病变,主要表现为成虫寄生于右心室,并可致心内膜,心肌及心外膜炎症性改变。     

Angiostrongylus cantonensis: Immunoblot analysis of the antigens recognized by rats

Following infection of rats with Angiostrongylus cantonensis, occurrence of anti-parasite antibody in the serum was determined with special reference to the antigens recognized by host IgG antibodies, using SDS-PAGE combined with an immunoblotting technique. Three saline extracts of digestive organ, reproductive organ and body wall, isolated from adult female A. cantonensis, were used as crude antigenic solutions. Then 7 to 49 days after infection, IgG antibodies directed predominantly against a single antigen, referred to as Ac-1 antigen, were detected. After 91 days or more, infected rats formed antibodies not only against the Ac-1 antigen, but also against a wide variety of other components with molecular weights in the range of 26000–220000 dalton. By using an antiserum against Ac-1 antigen as a probe, it was shown that the molecular weight and subunit structure, as well as the immunoelectrophoretic mobility, varied according to the organ from which the antigenic extract was prepared. The Ac-1 protein in the extract of the reproductive organ, one of the major sources of the Ac-1 antigen, showed the same electrophoretic mobility as -globulin. It has a molecular weight in the range of 100 000–200 000 dalton under both non-reducing and reducing conditions. Immunohistochemical studies, using the same antiserum and sectioned adult female worms, found Ac-1 antigen in the cytoplasm of oocytes at different stages of development, and in the lateral cord.     

Cellular immunity in Peyer''s patches of rats infected with Trichinella spiralis.

A rat model of Trichinella spiralis gut infection was used to observe the sequence of developing cellular immunity in Peyer's patches and other lymphoid tissues. Whereas cellular reactivity (lymphocyte blastogenesis) for worm antigens was evident in mesenteric lymph nodes draining the gastrointestinal tract within 3 days after infection, Peyer's patch lymphocytes developed maximal reactivity 2 to 3 weeks later at the same time as the spleen and other lymphoid tissues. Furthermore, the immune reactivity found in Peyer's patches was only transient. Thus, in this parasitic gut infection, the Peyer's patch lymphoid tissue does not appear to be the first site of cellular responsiveness but rather to acquire cellular reactivity only when other lymphoid elements in the infected host have also acquired similar antigen-induced reactivity.     

Effects of salmeterol on host resistance to Trichinella spiralis in rats.

Salmeterol is a long-acting beta2-adrenoreceptor agonist. The compound has previously been screened for immunotoxic potential in a repeated dose toxicity study in rats for 28 days. The total serum IgG levels were increased at dose levels of 2 and 10 mg/kg/day. Presently, salmeterol was studied in an immune function assay addressing the host resistance to Trichinella spiralis parasites. Rats were daily treated with salmeterol for 28 days at dose levels of 0, 2, 6 and 10 mg/kg/day. On day 29, the animals were infected with T. spiralis parasites. After six weeks, host resistance was examined. The numbers of T. spiralis muscle larvae in the tongue nor the inflammatory reactions around the encapsulated larvae were affected by salmeterol treatment. The yield of muscle larvae in the whole carcass was not changed either. The IgM, IgA and IgE antibody responses to T. spiralis were unaffected. Only at the highest dose level tested, the anti-T. spiralis IgG antibody response was decreased significantly. However, salmeterol's interference with the generation of anti-T. spiralis antibodies of the IgG subclass apparently did not adversely affect the resistance to infection.     

Phosphatidylethanolamine methylation in intestinal brush border membranes from rats resistant to Trichinella spiralis

Methylation of phospholipids is proposed as a mechanism to explain changes in properties of intestinal brush border membrane that coincide with development of immunity to the intraepithelial parasite, Trichinella spiralis. Methylation was measured by the incorporation of the [3H]methyl group from S-adenosyl-L-[3H]methyl methionine into phospholipids. At least two enzymatic components were detected that converted phosphatidylethanolamine to phosphatidylcholine. The first, designated methyltransferase I, catalyzed the formation of phosphatidylmonomethylethanolamine from phosphatidylethanolamine and had a low Km for S-adenosyl-L-methyl-methionine (5 microM). The second, designated methyltransferase II, which catalyzed the methylation of phosphatidylmonomethylethanolamine to phosphatidyldimethylethanolamine and phosphatidyldimethylethanolamine to phosphatidylcholine, had a high Km for S-adenosyl-L-methyl methionine (167 microM). Both enzymes had two pH optima, were most active at 37 degrees C and were Mg2+ dependent. A decrease in methylation activity was present in brush border membranes from rats immunized against T. spiralis. Although the synthesis of phosphatidylcholine was not significantly altered there was a substantial decrease in the formation of phosphatidylmonomethylethanolamine and phosphatidyldimethylethanolamine as compared with nonimmunized rats. Since phospholipid composition influences membrane fluidity and cell function, it is proposed that altered methylation activity may influence the characteristics of brush border membrane in the immune host.     

Immunocytochemical localization of antigens in adult worms ofTrichinella spiralis recognized by Fischer rats

We demonstrate the tissue localization, in adultTrichinella spiralis, of antigens recognized by Fischer rat sera at 32 weeks postinfection. Immunodominant antigens were located in a wide variety of tissues, including type 1 stichocyte granules, stichocyte cytoplasm, the canalicular tree, hemolymph, hypodermis, hypodermal glands, cord cytoplasm, intestinal-gland cell granules, membranous structures in the midgut epithelium, midgut-occupying substance, brush border, hindgut epithelial cytoplasm, hindgut cuticle, vaginal cuticle, epithelial cytoplasm of the female genital tract, microvilli and discrete areas of the ovum, embryo sheath, intersperm space, discrete areas in immature sperm, small granules and cup-shaped membrane structures of sperm, and exocrine granules in the seminal vesicle and ejaculatory duct. A small amount of antigen was located in the inner layers of the genital portion of the body cuticle. The precise localization of antigens in adult worms should form a basis for better analysis ofT. spiralis-related immunology.     

Rapid expulsion of Trichinella spiralis in suckling rats: mediation by monoclonal antibodies

Pups born to rats immunized with the excretory/secretory antigens (ESA) of Trichinella spiralis L1 larvae expressed rapid expulsion when challenged orally. Rat monoclonal antibodies specific for ESA were produced and tested for their specificity in Western blots, binding to intact larvae and protective capacity in suckling rats. Eight antibodies had apparently identical specificity in Western blots, each recognizing a polypeptide family that included three molecular weight species sized at 41,000-50,000 MW. These polypeptides formed a series of higher molecular weight aggregates that were also bound by the monoclonal antibodies. Four of eight antibodies were protective when serially transferred to suckling pups. Each protective antibody was able to bind to intact larvae. Antibodies of two subclasses, IgG1 and IgG2c, were strongly protective, delivering to pups the capacity to expel as much as 94% of the challenge dose.     

Immunoblot analysis of the circulating antigens occurring in serum of rats infected with Angiostrongylus cantonensis

Following the infection of rats with Angiostrongylus cantonensis, the occurrence and molecular features of circulating antigens (CAs) were analyzed, with special reference to their origin in worms, by SDS-PAGE combined with an immunoblotting technique. Antisera against the CAs were obtained by immunizing rats with sera from rats 35, 91, and 150 days postinfection. The antisera, referred to as anti-CA(35), anti-CA(91), and anti-CA(150), respectively, formed one or two precipitin lines when tested in Ouchterlony plates against extracts of digestive organs (DE) and of reproductive organs (RE) from adult female worms. When the anti-CA(35) was used as a blotting antibody under nonreducing conditions, a set of clearly spaced narrow bands with molecular weights (mol. wt.) in the range of 90000–180000 daltons developed only in the case of the DE. Besides the antigen(s), additional bands with mol.wt. of 115000 and 185000 daltons were revealed in the case of the RE when two other antisera were used. Immunoblot analysis of the immunoprecipitates, derived from anti-CA(150) and sera of infected rats, revealed the occurrence of two types of protein as the major CAs: one had a mol.wt. in the range of 140000–180000 daltons and was found in the serum 14 days postinfection, and the other, with a mol.wt. of 185000 daltons, was found in the serum 35–150 days postinfection. Immunohistochemical studies localized the CAs predominantly in the cytoplasm of both uterine eggs and maturing oocytes.     

Cross-resistance and cross-susceptibility between fluoroquinolone agents

The results of broth microdilution susceptibility tests with 750 bacterial isolates were used to directly compare six different fluoroquinolone antibioties (ciprofloxacin, enoxacin, fleroxacin, lomefloxacin, ofloxacin and temafloxacin). Against enteric bacilli, enoxacin and lomefloxacin were similar in their spectra of activity but they differed from the other drugs tested. Against the non-enteric gram-negative bacilli, ciprofloxacin differed from enoxacin and lomefloxacin: fleroxacin, ofloxacin and temafloxacin were nearly identical in their activity. For routine susceptibility tests, ofloxacin and fleroxacin were similar in activity and either drug could be used as a class representative for predicting susceptibility to the other fluoroquinolone agents. Occasionally, strains that are resistant to the class representative may be susceptible to other members of the fluoroquinolone class, but those that are susceptible to the class representative are rarely resistant to the other compounds.