Induction of T helper 1 response by immunization of BALB/c mice with the gene encoding the second subunit of Echinococcus granulosus antigen B (EgAgB8/2).

A pre-designed plasmid containing the gene encoding the second subunit of Echinococcus granulosus AgB8 (EgAgB8/2) was used to study the effect of the immunization route on the immune response in BALB/c mice. Mice were immunized with pDRIVEEgAgB8/2 or pDRIVE empty cassette using the intramuscular (i.m.), intranasal (i.n.) or the epidermal gene gun (g.g.) routes. Analysis of the antibody response and cytokine data revealed that gene immunization by the i.m. route induced a marked bias towards a T helper type 1 (Th1) immune response as characterized by high IFN-γ gene expression and a low IgG1/IgG2a reactivity index (R.I.) ratio of 0.04. The i.n. route showed a moderate IFN-γ expression but a higher IgG1/IgG2a R.I. ratio of 0.25 indicating a moderate Th1 response. In contrast, epidermal g.g. immunization induced a Th2 response characterized by high IL-4 expression and the highest IgG1/IgG2a R.I. ratio of 0.58. In conclusion, this study showed the advantage of genetic immunization using the i.m. route and i.n. over the epidermal g.g. routes in the induction of Th1 immunity in response to E. granulosus AgB gene immunization.

Tetracycline therapy targets intracellular bacteria in the filarial nematode Litomosoides sigmodontis and results in filarial infertility.

Intracellular bacteria have been described in several species of filarial nematodes, but their relationships with, and effects on, their nematode hosts have not previously been elucidated. In this study, intracellular bacteria were observed in tissues of the rodent parasite Litomosoides sigmodontis by transmission electron microscopy and by immunohistochemistry using antiendobacterial heat shock protein-60 antisera. Molecular phylogenetic analysis of the bacterial 16S ribosomal RNA gene, isolated by PCR, showed a close relationship to the rickettsial Wolbachia endobacteria of arthropods and to other filarial intracellular bacteria. The impact of tetracycline therapy of infected rodents on L. sigmodontis development was analyzed in order to understand the role(s) these bacteria might play in filarial biology. Tetracycline therapy, when initiated with L. sigmodontis infection, eliminated the bacteria and resulted in filarial growth retardation and infertility. If initiated after microfilarial development, treatment reduced filarial fertility. Treatment with antibiotics not affecting rickettsial bacteria did not inhibit filarial development. Acanthocheilonema viteae filariae were shown to lack intracellular bacteria and to be insensitive to tetracycline. These results suggest a mutualistic interaction between the intracellular bacteria and the filarial nematode. Investigation of such a mutualism in endobacteria-containing human filariae is warranted for a potential chemotherapeutic exploitation.

Immunocytogenetic effects of gonadotropin releasing hormone analogue: Triptorelin Pamoate (Decapeptyl) during in vitro fertilization treatment.

In this study, the immunocytogenetic effects of Decapeptyl (Triptorelin Pamoate) were assessed in the peripheral blood lymphocytes of females undergoing in vitro fertilization (IVF) treatment. Blood samples were taken from 34 females (23 treated and 11 controls), cultured and examined for sister chromatid exchanges (SCE) and cell replication index (CRI). The SCE frequency increased around ovulation time in the controls, and around the time of human chorionic gonadotropin administration in the IVF group. However, the SCE rate was significantly higher in the latter group. Furthermore, the white blood cells (WBC) count was significantly higher on the day of ovum pick up compared to the day preceding luteinizing hormone (LH) and follicle stimulating hormone (FSH) treatment. Similar observations were recorded with respect to phagocytic activity tested by nitroblue tetrazolium (NBT) assay. The nitric oxide production abilities of macrophages were not significantly changed in the LH, FSH-treated group relative to its control. Finally, the 50% complement hemolytic activity (CH50) assay results indicated that Decapeptyl lacks a significant potential to affect the complement system.

Lack of interferon-gamma confers impaired neutrophil granulocyte function and imparts prolonged survival of adult filarial worms in murine filariasis.

We investigated the role of IFN-gamma in host defense during murine filariasis. Using the fully permissive infection of BALB/c mice with the rodent filaria Litomosoides sigmodontis, we show that interferon (IFN)-gamma is essential for encapsulation of adult filarial worms in inflammatory nodules and for normal worm clearance. IFN-gamma knockout (KO) mice had only one third of the nodules of wild-type mice but displayed a more than twofold increase in worm burden and increased microfilaremia. Neutrophil granulocytes, but not macrophages or eosinophils, appear to directly control worm load and nodule formation. Neutrophils, which we showed earlier to be essential for the encapsulation process in the thoracic cavity, where the worms reside, were diminished at this location in IFN-gamma KO compared to wild-type mice; they also displayed strongly reduced chemotactic and phagocytic activity compared to neutrophils of controls. This argues for a distinct defect in neutrophil activation accounting for the low formation of inflammatory nodules. Tumor necrosis factor-alpha, a major neutrophil-activating cytokine expressed by macrophages in the thoracic cavity around the worms, was highly induced in wild-type but absent in KO mice. Diminished activation of neutrophils seems to be a general hallmark of IFN-gamma KO mice, since neutrophils from uninfected KO mice also showed a reduction in chemotactic and phagocytic activity when induced by casein. In conclusion, these data are the first to define an IFN-gamma-dependent immune effector mechanism in murine filarial infection, i.e. neutrophil-mediated control of the adult worm load.

Classification, follow-up and recurrence of hepatic cystic echinococcosis using ultrasound images.

Ultrasound image and morphological structure of hepatic cystic echinococcosis (CE) were analysed in 277 human cases (385 hepatic hydatid cysts). These included 65 CE cases from community surveys carried out between 1995 and 2000 in 3 countries (China, Mongolia and Jordan) and 212 cases from a clinical hospital survey from Xinjiang, China. A new simplified WHO ultrasound classification for human CE was assessed, and considered useful. It is proposed that type, size and number in particular need to be included in the ultrasound classification of hepatic CE. For comparative purposes 6 categories of type were classified in the study as Type 0 to Type 5 (T0-T5): T0, univesicular without pathognomonic signs; T1, univesicular with pathognomonic signs; T2, cysts with sagging or floating laminated membrane; T3, cysts containing daughter cysts; T4, solid mass or mixed cysts; and T5, cysts with partial or full calcifications. This differs from the WHO classification wherein Type T3 cysts (daughter cysts present) are considered a pathological stage to occur in general prior to the sagging or floating membrane (T2) stage. Recurrent hydatid cysts in the liver were also studied based on morphological structures observed directly from surgical intervention. Case follow-up over 1-5 years since endocystectomy in the community surveys indicated 10% (2/10) recurrence of cysts in the residual surgical cavity. Recurrent CE included 2 (2/4) cases after percutaneous treatment.

The Xid defect imparts susceptibility to experimental murine filariosis--association with a lack of antibody and IL-10 production by B cells in response to phosphorylcholine.

The pathways conferring immunity to filarial infections are not well known, in part because human pathogenic filariae do not develop a full infection cycle in laboratory mice. Using the permissive infection with Litomosoides sigmodontis in BALB/c mice, we have shown previously that worm development is controlled by CD4+ T cells and is inversely correlated with Th2 cytokine production. Here we analyzed the impact of the Xid immunodeficiency on murine filariosis, comparing the course of infection with L. sigmodontis in BALB/c and B1 cell-deficient BALB.Xid mice. In BALB.Xid mice, 2-3 times more adult worms and up to 10 times more microfilariae compared to BALB/c were observed to develop after infection with infective stage 3 larvae (L3). Parasite-specific Th2 cytokine production by cells from the thoracic cavity, the primary location of the parasites, was diminished significantly in BALB.Xid compared to BALB/c mice. In addition, BALB.Xid mice displayed a significantly lower production of antibodies and B cell-derived IL-10 in response to both L. sigmodontis antigen and phosphorylcholine, a molecule we found to be abundant on the surface of L3. Thus, the B cell-defect in BALB.Xid mice may account for susceptibility to murine filarial infection in two ways, i.e. by the lack of antibody to a dominant surface molecule of invading L3 and by less B cell-derived IL-10 resulting in lower parasite-driven Th2 cytokine production.

Human cystic echinococcosis in the West Bank of Palestine: surgical incidence and seroepidemiological study.

The surgical incidence of cystic echinococcosis (CE) due to Echinococcus granulosus was investigated in hospitals of the West Bank, Palestinian Authority between January 1990 and December 1997. Serum samples from school-children in Yata town, which showed the highest surgical incidence, were tested for anti-hydatid antibodies. A total of 390 surgically confirmed cases were recorded throughout the 8-year period, with an overall mean annual surgical incidence (MASI) of 3.1 per 100,000. A high MASI of 4.9, 5.0 and 5.1 per 100,000 was found in Hebron, Jericho and Bethlehem Governorates, respectively. Yata town, Hebron governorate, showed the highest MASI, at 16.8 per 100,000. The highest incidence was found in age groups 11-20 and 21-30 years, at 27.4% and 21.5% of the total number of cases. While there was no significant gender difference in the number of cases in the age groups of 20 years or less, the male to female case ratio was 1:3.2-4.1 in the older age groups. The liver was the most common site of hydatid cysts in 69.9% of cases. Lung cysts were predominant in younger age groups (20 years or less). The seropositivity for CE in the school-children of Yata was 2.4% and 2.1% using enzyme-linked immunosorbent assay and the indirect haemagglutination test, respectively. CE is a significant endemic disease throughout the West Bank. The disease is acquired early in life and is more prevalent among females than males. Behaviour and life-style favour the spread of the disease.

Infection of BALB/c mice with the filarial nematode Litomosoides sigmodontis: role of CD4+ T cells in controlling larval development.

Litomosoides sigmodontis is the only filaria which develops from infective larvae into adults in immunocompetent laboratory mice. Depletion of CD4+ T cells from infected BALB/c mice resulted in worm and microfilarial burdens significantly higher than those of infected controls. Th2 cytokines, eosinophilia, and immunoglobulin E, which were strongly induced in infected controls, were diminished in CD4-depleted mice.

A new mechanism for IL-5-dependent helminth control: neutrophil accumulation and neutrophil-mediated worm encapsulation in murine filariasis are abolished in the absence of IL-5.

IL-5 production and eosinophilia are features of helminth infections, but results concerning the role of IL-5 and eosinophils (EP) in worm control are contradictory. We describe here a novel, IL-5-dependent mechanism of helminth control in vivo, using a fully permissive murine filariasis model, i.e. infection of BALB/c mice with Litomosoides sigmodontis. Worm control was exerted by the formation of inflammatory nodules around adult filariae which initially remained alive but were eventually killed within several weeks. The cell population essential for inflammatory nodule formation was found to be neutrophils (NP) but not EP. Neutralization of IL-5 led to a failure of both EP and NP accumulation at the site of infection (i.e. the thoracic cavity), resulting in cessation of inflammatory nodule formation around worms and in their survival. The role of NP in this process was confirmed by treatment of mice with anti-granulocyte colony stimulating factor (G-CSF) which also resulted in a lack of inflammatory nodule formation and worm killing albeit in the presence of EP. Since IL-5, due to the absence of IL-5 receptors on NP, does not act on these cells directly, it was investigated if anti-IL-5 altered the production of NP-chemotactic cytokines. In anti-IL-5-treated mice, cytokines known to promote NP accumulation like tumor necrosis factor-alpha, G-CSF and KC (IL-8) were found to be strongly reduced, while NP-deactivating cytokines like IL-10 were increased. In conclusion, IL-5 constitutes a cytokine essential for NP-mediated worm control in filarial infection.

The seroprevalences of cystic echinococcosis, and the associated risk factors, in rural-agricultural, bedouin and semi-bedouin communities in Jordan.

An ELISA was used to determine the seroprevalence of cystic echinococcosis (CE), caused by Echinococcus granulosus, in representatives of the rural-agricultural, semi-bedouin and bedouin communities of Jordan. The knowledge, attitudes and practices (KAP) pertaining to the transmission of CE in such communities were also investigated. In the ELISA, serum samples from 2388 subjects were tested for IgG antibodies reacting with antigens in crude sheep hydatid fluid (CSHF). The rural-agricultural subjects were significantly more likely to be seropositive (11.4%) than the semi-bedouin (5.0%) or bedouin (3.7%), but male and female subjects were equally likely to be seropositive. Among the rural-agricultural and semi-bedouin subjects, those aged 11-20 years were most likely to be seropositive. Among the bedouin subjects, however, seroprevalence was highest for those aged 31-40 years. When the distribution of 59 seropositive subjects detected within 36 rural-agricultural households was investigated, 12 (33.3%) of the households were each found to have at least two seropositive members. Immunoblotting indicated that 27.1% of the seropositive rural-agricultural subjects showed immunoreactivity to at least one of the CSHF antigen-B components (of 8-12, 16 and 24 kDa). The living conditions, practices and lifestyles of the rural-agricultural, semi-bedouin and bedouin communities favour the spread of CE in Jordan and warrant an effective programme for the control of the disease.

IL-5 is essential for vaccine-induced protection and for resolution of primary infection in murine filariasis.

The pathways conferring immunity to human filariases are not well known, in part because human-pathogenic filariae do not complete a full life cycle in laboratory mice. We have used the only fully permissive infection of mice with filariae, i.e., infection of BALB/c mice with the rodent filarial nematode Litomosoides sigmodontis. Our previous results showed that worm development is inversely correlated with Th2 cytokine production and eosinophilia. The scope of the present study was to directly elucidate the role of interleukin-5 (IL-5) and eosinophils in controlling the development of L. sigmodonitis after vaccination and in primary infection. BALB/c mice immunized with irradiated third-stage larvae (L3) were confirmed to have elevated IL-5 levels as well as high subcutaneous eosinophilia and to attack and reduce incoming larvae within the first 2 days, resulting in 70% reduction of worm load. Treatment of vaccinated mice with anti-IL-5 antibody (TRFK-5) suppressed both blood and tissue eosinophilia and completely abolished protection. This demonstrates, for the first time in a fully permissive filarial infection, that IL-5 is essential for protection induced by irradiated L3 larvae. In contrast, in primary-infected mice, anti-IL-5 treatment did not modify filarial infection within the 1st month, most likely because during primary infection IL-5-dependent mechanisms such as subcutaneous eosinophilia are induced too late to disturb worm establishment. However, there is a role for IL-5 late in primary infection where neutrophil-dependent worm encapsulation is also under the control of IL-5.