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A Loukas

Publications and source records attributed to A Loukas.

At least 37 records · Page 2Linked to original sources
PubMed2000

Purification of a diagnostic, secreted cysteine protease-like protein from the hookworm Ancylostoma caninum.

The enteric infection of humans with the canine hookworm Ancylostoma caninum varies in its clinical presentation, ranging from asymptomatic to eosinophilic gastroenteritis requiring surgical intervention. Infections are not patent, but can be diagnosed immunologically by detecting antibodies to an immunodominant secreted hookworm protein termed Ac68. To characterise Ac68, we purified the native protein from A. caninum excretory/secretory products using size exclusion followed by anion exchange chromatography. The epitopes in the purified protein recognised by human infection sera were shown to be proteins and not carbohydrates. The N-terminal amino acid sequence of the purified Ac68 was determined and six of the 11 residues obtained were shared with a previously characterised cysteine protease of A. caninum, AcCP1.

Amino Acid Sequence↗
PubMed2000

A survey of genes expressed in adults of the human hookworm, Necator americanus.

Hookworms are gut-dwelling, blood-feeding nematodes that infect hundreds of millions of people, particularly in the tropics. As part of a program aiming to define novel drug targets and vaccine candidates for human parasitic nematodes, genes expressed in adults of the human hookworm Necator americanus were surveyed by the expressed sequence tag approach. In total 161 new hookworm genes were identified. For the majority of these, a function could be assigned by homology. The dataset includes proteases, protease inhibitors, a lipid binding protein, C-type lectins, an anti-bacterial factor, globins and other genes of interest from a drug or vaccine development viewpoint. Three different classes of small, secreted proteins were identified that may be involved in the host-parasite interaction, including potential potassium channel blocking peptides. One third of the genes were novel. These included highly expressed, secreted (glyco)proteins which may be part of the excretory-secretory products of these important pathogens. Of particular interest are a family of 9 genes with similarity to the immunomodulatory protein, neutrophil inhibitory factor, that may play a role in establishing an immunocompromised niche for this successful parasite.

Amino Acid Sequence↗
PubMed2000

Identification of a new C-type lectin, TES-70, secreted by infective larvae of Toxocara canis, which binds to host ligands.

Infective larvae of the dog roundworm Toxocara canis survive in the tissues of their hosts for extended periods in a state of developmental arrest, successfully evading immune destruction. This survival strategy is thought to be mediated by T. canis excretory/secretory (TES) products which downregulate or divert the immune response. We purified one of the major TES products, TES-70 and gained amino acid sequence from 4 tryptic peptides. These peptides were matched to a predicted protein from a cDNA that was isolated by expression screening a T. canis cDNA library with mouse anti-TES serum. The predicted protein (Tc-CTL-4) is similar to, but larger than, Tc-CTL-1, a 32-kDa C-type lectin secreted by T. canis larvae. Tc-CTL-4 has a signal peptide, 2 Cys-rich domains and a C-terminal calcium-dependent C-type lectin domain that shares sequence similarity with host immune cell receptors such as macrophage mannose receptor and CD23. The lectin domain was expressed in bacteria and antiserum to the purified recombinant protein was used to confirm that Tc-ctl-4 did encode the native TES-70 glycoprotein. TES-70 selectively bound to ligands on the surface of Madin-Darby Canine Kidney cells in vitro in a calcium-dependent manner, inhibitable by mammalian serum, indicating that a host glycan is the native ligand for this new parasite lectin.

Amino Acid Sequence↗
PubMed2000

The relation of personality to alcohol abuse/dependence in a high-risk sample.

The current study had two goals. The first goal was to test the mediational role of young adult personality in the relation between parental alcoholism and young adult alcoholism. The second was to examine the associations between personality and alcohol use motives and reasons to limit drinking in order to explore possible mechanisms by which personality may influence alcohol abuse/dependence. Multilevel modeling techniques were used to analyze data obtained from a community sample of young adult children of alcoholics and demographically matched controls. Results revealed that young adult neuroticism and agreeableness each, in part, mediated the effect of parental alcoholism on young adult alcoholism. Moreover, individuals high in neuloticism reported stronger coping motives to use alcohol, individuals low in agreeableness reported stronger coping motives and weaker upbringing reasons to limit drinking, and individuals low in conscientiousness reported stronger coping and enhancement motives to use alcohol, and weaker performance reasons to limit drinking.

Adolescent↗
PubMed1999

The organization of a galectin gene from Teladorsagia circumcincta.

Galectins are a family of soluble beta-galactoside-binding lectins that are conserved amongst a broad range of organisms. We have previously isolated cDNA clones coding for galectins from the sheep gastrointestinal nematode parasites Teladorsagia circumcincta, Haemonchus contortus and Trichostrongylus colubriformis, revealing a high level of identity between these molecules. This subsequent study reports the organization of the T. circumcincta Tci-gal-1 galectin gene. The coding region is broken into eight exons covering 6.6 kbp, with introns ranging in size from 55 base pairs (bp) to 2.8 kbp. Comparisons with recently reported galectin structures from Caenorhabditis elegans reveal strong architectural similarity between galectins from the parasitic and free-living nematodes, but this structure is not conserved in mammalian galectins.

Animals↗
PubMed1999

A hookworm allergen which strongly resembles calreticulin.

Immmoglobulin E-rich plasma from patients from Papua New Guinea infected with Necator americanus has been used to probe an adult N. americanus cDNA library for the presence of hookworm allergens. Using this approach, one hookworm allergen has been identified as calreticulin, which was subsequently expressed in Escherichia coli. Little serological cross reactivity was seen between the recombinant calreticulins of this hookworm and its host. Prospective roles for hookworm calreticulin in the host-parasite relationship are discussed in depth.

Allergens↗
PubMed1999

Identification of abundantly expressed novel and conserved genes from the infective larval stage of Toxocara canis by an expressed sequence tag strategy.

Larvae of Toxocara canis, a nematode parasite of dogs, infect humans, causing visceral and ocular larva migrans. In noncanid hosts, larvae neither grow nor differentiate but endure in a state of arrested development. Reasoning that parasite protein production is orientated to immune evasion, we undertook a random sequencing project from a larval cDNA library to characterize the most highly expressed transcripts. In all, 266 clones were sequenced, most from both 3' and 5' ends, and similarity searches against GenBank protein and dbEST nucleotide databases were conducted. Cluster analyses showed that 128 distinct gene products had been found, all but 3 of which represented newly identified genes. Ninety-five genes were represented by a single clone, but seven transcripts were present at high frequencies, each composing >2% of all clones sequenced. These high-abundance transcripts include a mucin and a C-type lectin, which are both major excretory-secretory antigens released by parasites. Four highly expressed novel gene transcripts, termed ant (abundant novel transcript) genes, were found. Together, these four genes comprised 18% of all cDNA clones isolated, but no similar sequences occur in the Caenorhabditis elegans genome. While the coding regions of the four genes are dissimilar, their 3' untranslated tracts have significant homology in nucleotide sequence. The discovery of these abundant, parasite-specific genes of newly identified lectins and mucins, as well as a range of conserved and novel proteins, provides defined candidates for future analysis of the molecular basis of immune evasion by T. canis.

3' Untranslated Regions↗
PubMed1998

Characterisation of Tc-cpl-1, a cathepsin L-like cysteine protease from Toxocara canis infective larvae.

Cysteine proteases play vital biological roles in both intracellular and extracellular environments. A cysteine protease migrating at 30 kDa was identified in somatic extracts of Toxocara canis larvae (TEX), by its binding to the biotinylated inhibitor Phe-Ala-CH2F. TEX proteases readily cleaved the cathepsin L- and B-specific peptide substrate Z-Phe-Arg-AMC and to a lesser extent, the cathepsin B-specific peptide Z-Arg-Arg-AMC. Excretory/secretory (TES) products of T. canis larvae did not cleave either substrate. Partial sequence encoding the 5' end of a cysteine protease cDNA from infective T. canis larvae was then obtained from an expressed sequence tag (EST) project. The entire cDNA (termed Tc-cpl-1) was subsequently sequenced and found to encode a preproenzyme similar to cathepsin L-like proteases (identities between 36 and 69%), the closest homologues being two predicted proteins from Caenorhabditis elegans cosmids, a cathepsin L-like enzyme from Brugia pahangi and a range of parasite and plant papain-like proteases. Sequence alignment with homologues of known secondary structure indicated several charged residues in the S1 and S2 subsites involved in determining substrate specificity. Some of these are shared with human cathepsin B, including Glu 205 (papain numbering), known to permit cleavage of Arg-Arg peptide bonds. The recombinant protease (rTc-CPL-1) was expressed in bacteria for immunisation of mice and the subsequent antiserum shown to specifically react with the 30 kDa native protease in TEX. Sera from mice infected with the parasite also contained antibodies to rTc-CPL-1 as did sera from nine patients with proven toxocariasis; control sera did not. Larger scale studies are underway to investigate the efficacy of rTc-CPL-1 as a diagnostic antigen for human toxocariasis, the current test for which relies on whole excretory/secretory antigens of cultured parasites.

Amino Acid Sequence↗
PubMed1998

Cloning and characterisation of a prohibitin gene from infective larvae of the parasitic nematode Toxocara canis.

Infective larvae of the parasitic nematode Toxocara canis express an mRNA (Tc-pro-1) encoding a predicted protein that shares significant homology with prohibitin, a protein involved in inhibition of cell proliferation - The closest homologues of Tc-pro-1 include an expressed sequence tag (EST) from Caenorhabditis elegans and Drosophila L2Cc, a protein thought to be essential for larval development and moulting. Other homologues include prohibitin from rat and human and an EST from Saccharomyces cerevisiae. Parasite life cycles generally include periods of developmental arrest, which in the larvae of T. canis may persist for many years without loss of metabolic activity. This report of the first full-length gene encoding prohibitin from a parasitic nematode raises interesting suggestions about the potential role of prohibitin in diapause and in the regulation of moulting in development.

Amino Acid Sequence↗
PubMed1996

A distinctive aphthous ileitis linked to Ancylostoma caninum.

Ileal ulceration is diagnosed commonly at colonoscopy in northeastern Australia. This observation has occurred contemporaneously with the description of a painful enteritis caused through supposedly rare infections by Ancylostoma caninum, the cosmopolitan dog hookworm. In this study, we describe the morphology and prevalence of ileal ulcers and investigate their cause. There were 182 patients with a distinctive aphthous ileitis highlighted by haemorrhage into the tips of surrounding villi. The annual rate reached 4.7% of ileal examinations. Eosinophilic enteritis was the characteristic, but inconsistent, histological appearance. Ulceration was linked to infection by A. caninum through serological testing and, in three patients, by the retrieval of single hookworms. Affected patients were also more likely to own a dog (53%) than the total colonoscopic population (43%; P < 0.02). The monthly rates of diagnosis were related to climate with fewest recordings during winter. Pain was the chief indication for colonoscopy (49%), proportionally higher than in controls (28%; P < 0.0001). Patients with ulcers had increased blood eosinophil, immunoglobulin E and serological values. These results were higher still in the subset with pain compared with the rest. Most patients, however, had colonoscopy performed for unrelated reasons, such as bowel cancer surveillance, and were identified as having subclinical ulceration (rate 3.2%). The findings establish a distinctive aphthous ileitis characterized by tissue eosinophilia. The cause has been tentatively linked to canine hookworms. It is proposed that infection in this geographic region is common and only rarely does a painful enteritis develop as a consequence of hypersensitization.

Ancylostoma↗
PubMed1996

Immunoglobulin G subclass antibodies against excretory/secretory antigens of Ancylostoma caninum in human enteric infections.

Most patients with proven or suspected enteric infection with the common hookworm of dogs, Ancylostoma caninum, produce immunoglobulin G (IgG) and IgE antibodies to an immunodominant excretory/secretory antigen (Ac68) of the parasite. These antibodies were detected in both enzyme-linked immunosorbent assay (ELISA) and Western blots; the Western blot to detect IgG antibodies to Ac68 was the most specific and sensitive. The subclasses of IgG of the antibody response to the parasite were analyzed using Western blots with anti-IgG subclass-specific monoclonal antibodies as marker systems in an attempt to further improve the specificity of the assay. Eight patients with confirmed enteric infections with A. caninum (positive controls) were tested; six had antibodies in all IgG subclasses against Ac68. Twenty sera from patients with suspected enteric infection with A. caninum (manifested as eosinophilic enteritis or unexplained abdominal pain with peripheral eosinophilia) were tested; 16 had total IgG antibodies to Ac68, while IgG1, IgG2, IgG3, and IgG4 responses were found in 11, 10, 9, and 12 of these sera, respectively. Small numbers of sera from groups of patients infected with other helminths and from healthy blood donors had various combinations of IgG, IgG1, IgG2, and IgG3 antibodies to Ac68, but none of these sera had IgG4 antibodies to Ac68. Sera from all nine patients with human hookworm infection had IgG, IgG1, IgG2, and IgG3 antibodies to Ac68 and eight of the nine were also positive for IgG4 antibodies. These results indicate the Western blot to detect IgG4 antibodies to Ac68 is the most reliable immunodiagnostic test yet described for enteric infection with A. caninum, although this test does not discriminate between infections with human and canine hookworms.

Ancylostoma↗
PubMed1995

Eosinophilic enteritis in northeastern Australia. Pathology, association with Ancylostoma caninum, and implications.

While eosinophilic gastroenteritis is considered a rare condition, eosinophilic enteritis without gastric involvement is quite common in northeastern Australia. We present 79 patients with biopsy-proven eosinophilic enteritis, 70 seen since 1987. In 10 patients, eosinophilic enteritis was associated with infection by single, sexually immature, adult hookworms, most positively identified as the common dog hookworm Ancylostoma caninum. An additional 22 patients (of 34 tested) had serological evidence of A. caninum exposure. The essential pathology, i.e., edema and eosinophilic infiltration of the gut wall, ascites, and regional lymphadenopathy, was identical to that seen in eosinophilic gastroenteritis. Additional, more specific features included pathological reactions centered on attached worms, mucosal alterations and ulcers considered to be hookworm bite sites, and submucosal and lymph node granulomas with central eosinophil degranulation and degradation products. Since A. caninum has an almost worldwide distribution, it is probable that A. caninum-induced eosinophilic enteritis occurs outside Australia. We show that the worm is easily overlooked in pathological specimens and that care is required to preserve worms intact for specific parasitological identification. The clinical and pathological features were similar to those seen in another human enteric helminthic zoonosis, anisakiasis. The possibility that there are yet other undiscovered intestinal zoonoses remains.

Adolescent↗
PubMed1994

Human enteric infection with canine hookworms.

OBJECTIVE: To describe a zoonotic ancylostomiasis (canine), acquired from domestic pets by patients living in developed, urban communities. DESIGN: An 8-year, retrospective case study. SETTING: A clinical gastroenterologic practice in Townsville and a university parasitology department in Brisbane, Australia. PATIENTS: Nine patients, each with enteric hookworm infection diagnosed by finding a single organism in situ; five were treated by us, and the rest were referred to us for parasite identification. MEASUREMENTS: Clinical and demographic data, complete blood examinations, total serum immunoglobulin E assay, and serologic testing with enzyme-linked immunosorbent assay and Western blot using excretory-secretory antigens of Ancylostoma caninum. Gut biopsy specimens were examined histologically, and hookworms were identified using morphologic criteria. RESULTS: The infections in three of the patients were diagnosed during the initial 6 years and six in the last 2 years. All owned a dog and described activity potentially exposing them to infection with canine hookworm larvae. Three patients had a laparotomy for acute abdominal pain, and six had colonoscopies (five with pain and one without symptoms). Six of the nine had blood eosinophilia (mean, 0.97 x 10(9)/L), and five of eight had elevated immunoglobulin E levels (mean level, 756 micrograms/L); six of eight had eosinophilic inflammation of the gut. In six patients, the worm was identified as A. caninum, whereas in three, damage to the specimen did not allow specific identification; however, they were unlikely to be human parasite species. Although all parasites were in the adult stage, none were sexually mature. Positive serologic findings in seven of the eight patients tested confirmed presence of antibody to the parasite. CONCLUSIONS: Human enteric infections with A. caninum are being diagnosed more frequently in northeastern Australia. Although infection may be subclinical, the chief symptom is abdominal pain, sometimes sudden and severe. The pathologic finding is focal or diffuse eosinophilic inflammation caused by a type 1 hypersensitivity response to secreted antigens. Infection by sexually immature worms is scant and nonpatent, indicating poor adaptation to the human host. Serologic testing assists in identification of occult infection. Advanced hygiene and sanitation afford little protection because the parasite reservoir is a large and growing pool of infected domestic pets.

Abdominal Pain↗
PubMed1994

Occult enteric infection by Ancylostoma caninum: a previously unrecognized zoonosis.

BACKGROUND/AIMS: Human disease caused by the canine hookworm Ancylostoma caninum and a high incidence of eosinophilic enteritis have been reported from northern Queensland, Australia. The aim of this study was to evaluate patients with unexplained abdominal pain and a possible association with cryptic infections by A. caninum. METHODS: The clinical and demographic features of patients from this region with eosinophilic enteritis (group A1, n = 42), obscure abdominal pain associated with (group A2, n = 105) and without (group A3, n = 84) blood eosinophilia were reviewed and sera were tested against A. caninum excretory-secretory antigens by enzyme-linked immunosorbent assay (ELISA) and Western blot. Four additional patients, two with confirmed A. caninum, had hookworm infection. RESULTS: The level of dog ownership in these four groups was 79%-100%, higher than in the local population (P < 0.001). The ELISA tested positive in 71% of A1, 67% of A2, and 30% of A3, versus 8% in controls (P < 0.002). All cases tested were positive on Western blot versus 10% of controls (P < 0.0001). The ELISA values increased with chronicity and decreased during convalescence. CONCLUSIONS: We conclude occult human A. caninum infections are common and are characterized by eosinophilic enteritis and obscure abdominal pain with or without blood eosinophilia. The diagnosis can be confirmed by serology.

Adolescent↗
PubMed1994

Immunologic incrimination of Ancylostoma caninum as a human enteric pathogen.

In northeastern Australia, the incidence of human eosinophilic enteritis (EE) and unexplained abdominal pain with peripheral blood eosinophilia (PE) appear to be exceptionally high. Because adults of the common dog hookworm, Ancylostoma caninum, were found in situ in several cases, we compared specific antibody responses in patients with EE or PE with those of individuals from various control groups. Sera were obtained from three patients with confirmed dog hookworm infection, 25 patients with EE/PE, 42 with other diagnosed gastrointestinal disorders, eight with human hookworm infection, 27 with other diagnosed parasitic infections, and 100 blood donors from the State of Tasmania, where A. caninum does not occur. They were analyzed by enzyme-linked immunosorbent assay (ELISA) and Western blot for IgG and IgE antibodies to excretory-secretory (ES) antigens from adult A. caninum. In the ELISA, sera from 88% of EE/PE patients were positive for IgG and IgE antibodies to ES antigen. All eight patients infected with human hookworm (identified as A. duodenale in three) were also seropositive, while most of those with other parasitic infections, as well the blood donors, those with diagnosed gastrointestinal diseases, and the control subjects, were seronegative. In Western blots, sera from 92% of EE/PE patients demonstrated IgG and IgE antibodies to a component of ES antigen with a molecular weight of approximately 68 kD (Ac68), as did all sera from cases infected with human hookworms. Among the patients with other gastrointestinal disorders and parasitic infections, four of 42 (10%) and three of 27 (11%), respectively, reacted positively.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗