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P Upcroft

Publications and source records attributed to P Upcroft.

At least 19 recordsLinked to original sources
PubMed2003

Giardia duodenalis trophozoites isolated from a parrot (Cacatua galerita) colonize the small intestinal tracts of domestic kittens and lambs.

This study examines the ability of Giardia duodenalis trophozoites, isolated from a wild bird, to colonize the intestinal tracts of companion animals (kittens) and domestic ruminants (lambs). Trophozoites colonized the intestinal tracts of intraduodenally inoculated animals as demonstrated by increasing parasite burdens within the duodenum and jejunum and by fecal passage of cysts within 4 days post-inoculation. The pathogenesis of the trophozoites was further investigated in kittens. In these animals, infection significantly reduced jejunal brush border microvillous length and density, which resulted in a loss of overall epithelial brush border surface area. This injury was associated with the production of diarrhea in four of five infected kittens. These findings indicate that some bird species may carry G. duodenalis that represent a possible health threat to companion animals and livestock. Our results describe the first successful colonization of avian-derived G. duodenalis trophozoites in the small intestines of domestic kittens and lambs.

Animals↗
PubMed2001

Drug susceptibility testing of anaerobic protozoa.

A simple technique for routine, reproducible global surveillance of the drug susceptibility status of the anaerobic protozoa Trichomonas, Entamoeba, and Giardia is described. Data collected using this technique can be readily compared among different laboratories and with previously reported data. The technique employs a commercially available sachet and bag system to generate a low-oxygen environment and log(2) drug dilutions in microtiter plates, which can be monitored without aerobic exposure, to assay drug-resistant laboratory lines and clinically resistant isolates. MICs (after 2 days) of 3.2 and 25 microM indicated metronidazole-sensitive and highly clinically resistant isolates of T. vaginalis in anaerobic assays, respectively. The aerobic MICs were 25 and >200 microM. MICs (1 day) of 12.5 to 25 microM were found for axenic lines of E. histolytica, and MICs for G. duodenalis (3 days) ranged from 6.3 microM for metronidazole-sensitive isolates to 50 microM for laboratory metronidazole-resistant lines. This technique should encourage more extensive monitoring of drug resistance in these organisms.

Albendazole↗
PubMed2001

Drug targets and mechanisms of resistance in the anaerobic protozoa.

The anaerobic protozoa Giardia duodenalis, Trichomonas vaginalis, and Entamoeba histolytica infect up to a billion people each year. G. duodenalis and E. histolytica are primarily pathogens of the intestinal tract, although E. histolytica can form abscesses and invade other organs, where it can be fatal if left untreated. T. vaginalis infection is a sexually transmitted infection causing vaginitis and acute inflammatory disease of the genital mucosa. T. vaginalis has also been reported in the urinary tract, fallopian tubes, and pelvis and can cause pneumonia, bronchitis, and oral lesions. Respiratory infections can be acquired perinatally. T. vaginalis infections have been associated with preterm delivery, low birth weight, and increased mortality as well as predisposing to human immunodeficiency virus infection, AIDS, and cervical cancer. All three organisms lack mitochondria and are susceptible to the nitroimidazole metronidazole because of similar low-redox-potential anaerobic metabolic pathways. Resistance to metronidazole and other drugs has been observed clinically and in the laboratory. Laboratory studies have identified the enzyme that activates metronidazole, pyruvate:ferredoxin oxidoreductase, to its nitroso form and distinct mechanisms of decreasing drug susceptibility that are induced in each organism. Although the nitroimidazoles have been the drug family of choice for treating the anaerobic protozoa, G. duodenalis is less susceptible to other antiparasitic drugs, such as furazolidone, albendazole, and quinacrine. Resistance has been demonstrated for each agent, and the mechanism of resistance has been investigated. Metronidazole resistance in T. vaginalis is well documented, and the principal mechanisms have been defined. Bypass metabolism, such as alternative oxidoreductases, have been discovered in both organisms. Aerobic versus anaerobic resistance in T. vaginalis is discussed. Mechanisms of metronidazole resistance in E. histolytica have recently been investigated using laboratory-induced resistant isolates. Instead of downregulation of the pyruvate:ferredoxin oxidoreductase and ferredoxin pathway as seen in G. duodenalis and T. vaginalis, E. histolytica induces oxidative stress mechanisms, including superoxide dismutase and peroxiredoxin. The review examines the value of investigating both clinical and laboratory-induced syngeneic drug-resistant isolates and dissection of the complementary data obtained. Comparison of resistance mechanisms in anaerobic bacteria and the parasitic protozoa is discussed as well as the value of studies of the epidemiology of resistance.

Anaerobiosis↗
PubMed2001

Orally administered Giardia duodenalis extracts enhance an antigen-specific antibody response.

We have identified novel adjuvant activity in specific cytosol fractions from trophozoites of Giardia isolate BRIS/95/HEPU/2041 (J. A. Upcroft, P. A. McDonnell, and P. Upcroft, Parasitol. Today, 14:281-284, 1998). Adjuvant activity was demonstrated in the systemic and mucosal compartments when Giardia extract was coadministered orally with antigen to mice. Enhanced antigen-specific serum antibody responses were demonstrated by enzyme-linked immunosorbent assay to be comparable to those generated by the "gold standard," mucosal adjuvant cholera toxin. A source of adjuvant activity was localized to the cytosolic component of the parasite. Fractionation of the cytosol produced fraction pools, some of which, when coadministered with antigen, stimulated an enhanced antigen-specific serum response. The toxic component of conventional mucosal adjuvants is associated with adjuvant activity; therefore, in a similar way, the toxin-like attributes of BRIS/95/HEPU/2041 may be responsible for its adjuvanticity. Complete characterization of the adjuvant is under way.

Adjuvants, Immunologic↗
PubMed2000

Immune and pathophysiological responses to different strains of Giardia duodenalis in neonatal mice.

Numerous studies have demonstrated various strain differences between Giardia isolates, but little is known about the immunology and pathogenesis of infections. This study aimed to compare host responses to strains of Giardi duodenalis differing in levels of virulence and pathogenicity and, by doing so, elucidate the mechanisms via which pathogenic strains establish infections. Marked differences were found in the infection dynamics, histopathological responses and serum antibody responses of neonatal mice infected with either G. duodenalis strain BRIS/83/HEPU/106 (isolated from a human) or BRIS/95/HEPU/2041 (isolated from a sulphur-crested cockatoo, Cacatua galerita). Infections with the bird strain were more intense (6.7-times greater) and persisted longer (by 14days) than infections with the human strain. The bird strain was more pathogenic and caused greater pathophysiological alteration to the gut mucosa, including increased villous atrophy, hyperplasia of goblet cells and vacuolated epithelial cells. Mice infected with the bird strain produced less serum anti-Giardia IgA and IgM, but more total (non-specific) serum IgA than those infected with the human strain of Giardia. This suggests that avian G. duodenalis strains are infective for mammalian hosts and may contribute to zoonotic infections. Furthermore, infection of mice with BRIS/95/HEPU/2041 serves as a good experimental model to provide further insight into the mechanisms via which G. duodenalis causes disease.

Animals↗
PubMed1999

Alternative 2-keto acid oxidoreductase activities in Trichomonas vaginalis.

We have induced high levels of resistance to metronidazole (1 mM or 170 microg ml(-1)) in two different strains of Trichomonas vaginalis (BRIS/92/STDL/F1623 and BRIS/92/STDL/B7708) and have used one strain to identify two alternative T. vaginalis 2-keto acid oxidoreductases (KOR) both of which are distinct from the already characterised pyruvate:ferredoxin oxidoreductase (PFOR). Unlike the characterised PFOR which is severely down-regulated in metronidazole-resistant parasites, both of the alternative KORs are fully active in metronidazole-resistant T. vaginalis. The first, KORI, localized in all membrane fractions but predominantly in the hydrogenosome fraction, is soluble in Triton X-100 and the second, KOR2, is extractable in 1 M acetate from membrane fractions of metronidazole-resistant parasites. PFOR and both KORI and KOR2 use a broad range of 2-keto acids as substrates (pyruvate, alpha-ketobutyrate, alpha-ketomalonate), including the deaminated forms of aromatic amino acids (indolepyruvate and phenylpyruvate). However, unlike PFOR neither KORI or KOR2 was able to use oz-ketoglutarate. Deaminated forms of branched chain amino acids (alpha-ketoisovalerate) were not substrates for T. vaginalis KORs. Since KOR I and KOR2 do not apparently donate electrons to ferredoxin, and are not down-regulated in metronidazole-resistant parasites, we propose that KORI and KOR2 provide metronidazole-resistant parasites with an alternative energy production pathway(s) which circumvents metronidazole activation.

Animals↗
PubMed1999

Efficacy of new 5-nitroimidazoles against metronidazole-susceptible and -resistant Giardia, Trichomonas, and Entamoeba spp.

The efficacies of 12 5-nitroimidazole compounds and 1 previously described lactam-substituted nitroimidazole with antiparasitic activity, synthesized via SRN1 and subsequent reactions, were assayed against the protozoan parasites Giardia duodenalis, Trichomonas vaginalis, and Entamoeba histolytica. Two metronidazole-sensitive lines and two metronidazole-resistant lines of Giardia and one line each of metronidazole-sensitive and -resistant Trichomonas were tested. All except one of the compounds were as effective or more effective than metronidazole against Giardia and Trichomonas, but none was as effective overall as the previously described 2-lactam-substituted 5-nitroimidazole. None of the compounds was markedly more effective than metronidazole against Entamoeba. Significant cross-resistance between most of the drugs tested and metronidazole was evident among metronidazole-resistant lines of Giardia and Trichomonas. However, some drugs were lethal to metronidazole-resistant Giardia and had minimum lethal concentrations similar to that of metronidazole for drug-susceptible parasites. This study emphasizes the potential in developing new nitroimidazole drugs which are more effective than metronidazole and which may prove to be useful clinical alternatives to metronidazole.

Animals↗
PubMed1998

A dynein heavy chain homologue gene in Hexamita inflata.

A gene encoding an unusually small dynein heavy chain homologue, hDYHH, was cloned from the genome of a free-living diplomonad, Hexamita inflata (Hi). The open reading frame (ORF) of hDYHH is 867bp and encodes a polypeptide of 289 amino acids (aa), hDYHH. hDYHH is homologous to the region around the third P-loop ATP-binding site of several dynein heavy chain polypeptides that are around 4000aa. Northern blot analysis showed that hDYHH is expressed in vivo and that the mRNA length (approximately 1.8kb) is consistent with the gene length (1.67kb). Southern blot analysis indicated that there are hDYHH homologues within the Hi genome, possibly including a longer dynein heavy chain gene. An hDYHH homologue was also identified in Hexamita pusilla (Hp). hDYHH is the first full-length protein-encoding gene cloned from Hexamita.

Amino Acid Sequence↗
PubMed1998

My favorite cell: Giardia.

The gut protozoan parasite, Giardia duodenalis, is the best characterized example of the most ancient eukaryotes, which are anaerobic and appear to be primitively amitochondrial. Apart from its obvious medical importance, Giardia is fascinating in its own right. Its prokaryotic-like anaerobic metabolism renders it selectively sensitive to some bacterial drugs, especially the nitroimidazoles, which are activated to form toxic radicals. Other features, including an enzyme that reduces oxygen directly to water, cysteine as the keeper of redox balance, a plasmid, and toxin-like genes are also distinctly prokaryotic-like. But, unlike prokaryotes, Giardia has a sophisticated, highly developed cytoskeleton, bounded nuclei, linear chromosomes capped with telomeric repeats, and telomere positional regulation of gene expression.

Animals↗
PubMed1998

Anaerobic bacterial metabolism in the ancient eukaryote Giardia duodenalis.

The protozoan parasite, Giardia duodenalis, shares many metabolic and genetic attributes of the bacteria, including fermentative energy metabolism which relies heavily on pyrophosphate rather than adenosine triphosphate and as a result contains two typically bacterial glycolytic enzymes which are pyrophosphate dependent. Pyruvate decarboxylation and subsequent electron transport to as yet unidentified anaerobic electron acceptors relies on a eubacterial-like pyruvate:ferredoxin oxidoreductase and an archaebacterial/eubacterial-like ferredoxin. The presence of another 2-ketoacid oxidoreductase (with a preference for alpha-ketobutyrate) and multiple ferredoxins in Giardia is also a trait shared with the anaerobic bacteria. Giardia pyruvate:ferredoxin oxidoreductase is distinct from the pyruvate dehydrogenase multienzyme complex invariably found in mitochondria. This is consistent with a lack of mitochondria, citric acid cycle, oxidative phosphorylation and glutathione in Giardia. Giardia duodenalis actively consumes oxygen and yet lacks the conventional mechanisms of oxidative stress management, including superoxide dismutase, catalase, peroxidase, and glutathione cycling, which are present in most eukaryotes. In their place Giardia contains a prokaryotic H2O-producing NADH oxidase, a membrane-associated NADH peroxidase, a broad-range prokaryotic thioredoxin reductase-like disulphide reductase and the low molecular weight thiols, cysteine, thioglycolate, sulphite and coenzyme A. NADH oxidase is a major component of the electron transport pathway of Giardia which, in conjunction with disulphide reductase, protects oxygen-labile proteins such as ferredoxin and pyruvate:ferredoxin oxidoreductase against oxidative stress by maintaining a reduced intracellular environment. As the terminal oxidase, NADH oxidase provides a means of removing excess H+, thereby enabling continued pyruvate decarboxylation and the resultant production of acetate and adenosine triphosphate. A further example of the bacterial-like metabolism of Giardia is the utilisation of the amino acid arginine as an energy source. Giardia contain the arginine dihydrolase pathway, which occurs in a number of anaerobic prokaryotes, but not in other eukaryotes apart from trichomonads and Chlamydomonas reinhardtii. The pathway includes substrate level phosphorylation and is sufficiently active to make a major contribution to adenosine triphosphate production. Two enzymes of the pathway, arginine deiminase and carbamate kinase, are rare in eukaryotes and do not occur in higher animals. Arginine is transported into the trophozoite via a bacterial-like arginine:ornithine antiport. Together these metabolic pathways in Giardia provide a wide range of potential drug targets for future consideration.

Amino Acids↗
PubMed1998

Virulent Avian Giardia duodenalis Pathogenic for Mice.

Early in 1995, a sulphur-crested cockatoo captured in the wild died along with several other cage mates, apparently of an overwhelming, acute infection of Giardia. Trophozoites isolated from the dead bird and established in traditional Giardia axenic medium were infective to mice and established chronic infections associated with weight gain impairment. Genetically and morphologically, the Giardia isolated from the bird belonged to the duodenalis group. Here, Jacqui Upcroft, Ann McDonnell and Peter Upcroft present data on pathogenic avian Giardia with he potential to contaminate watersheds and discuss the implications.

Journal Article↗
PubMed1998

Drug resistance in Giardia: clinical versus laboratory isolates.

The advantages and limitations of determining mechanisms of drug resistance in Giardia duodenalis laboratory isolates, which have been generated in a number of ways, is weighed against the difficulty of analysing mechanisms in clinical isolates with a large diversity of genetic and expression capabilities. Using isogenic strains to follow changes in enzyme regulation involved in drug resistance, we have been able to assess the full capability of the parasite in developing drug resistance mechanisms. The complementarity of the two approaches, clinical versus laboratory induced drug resistance, and continuing comparison with other organisms, particularly the anaerobic bacteria with which Giardia has strong affiliations, is emphasized. These considerations lead to the study of the population genetics of drug resistance, and strategies critical for rational drug usage, design and therapy.

Journal Article↗
PubMed1997

Lethal Giardia from a wild-caught sulphur-crested cockatoo (Cacatua galerita) established in vitro chronically infects mice.

An axenic culture of Giardia was established from a sample of infected intestine obtained following autopsy of a sulphur-crested cockatoo (Cacatua galerita). The cockatoo recently captured in the wild and with good muscle tone died along with several other cage mates, apparently of an overwhelming, acute infection of Giardia. Trophozoites which established in the traditional, axenic Giardia medium (TYI-S-33 with supplementary bile) were morphologically identical to G. duodenalis. When outbred Quackenbush Swiss neonatal mice were infected with trophozoites a chronic infection was established and parasites were still present at 38 days post-inoculation. Weight gain by infected mice was reduced by 20%, thus mimicking failure-to-thrive syndrome in children, and maximum parasite load was more than 3-fold higher in comparison with other G. duodenalis strains. Analysis of the electrophoretic karyotype, rDNA and hybridization studies together with Giemsa- and trichrome-stained samples, and scanning electron microscopy indicated that the bird-derived Giardia belonged to the duodenalis group. This is the first report of infection of mammals with Giardia isolated from a bird. These data may have potentially serious implications for contamination of watersheds and establishment of zoonotic infections.

Animals↗