|
1
|
Nagel R, Traub RJ, Kwan MMS, Bielefeldt-Ohmann H. Blastocystis specific serum immunoglobulin in patients with irritable bowel syndrome (IBS) versus healthy controls. Parasit Vectors 2015; 8:453. [PMID: 26373392 PMCID: PMC4572630 DOI: 10.1186/s13071-015-1069-x] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2015] [Accepted: 09/03/2015] [Indexed: 12/14/2022] Open
Abstract
Background Blastocystis species are common enteric human parasites and carriage has been linked to Irritable Bowel Syndrome (IBS), particularly diarrhoea-predominant IBS. The spectrum of immune reactivity to Blastocystis proteins has been reported previously in symptomatic patients. We investigated differences in serum immunoglobulin profiles between patients with IBS, both positive and negative for Blastocystis carriage, and healthy controls (HC). Methods Forty diarrhoea-predominant IBS patients (26 patients positive for Blastocystis sp., 14 negative patients) and forty HC (24 positive, 16 Blastocystis-negative) were enrolled. Age, gender, ethnicity and serum immunoglobulin A (IgA) levels were recorded and faecal specimens were analysed using smear, culture and polymerase chain reaction amplification of ribosomal DNA. Sera were tested in Western blots and the reactivities compared to known targets using monoclonal antibodies Blastofluor® (Blastocystis specific antibody), MAb1D5 (cytopathicto Blastocystis cells), anti-promatrix metalloprotease-9 (anti-MMP-9) and SDS-PAGE zymograms. Results Levels of serum IgA were significantly lower in Blastocystis carriers (p < 0.001) but had no relationship to symptoms. Western blots demonstrated serum IgG antibodies specific for Blastocystis proteins of 17,27,37,50,60-65, 75–90, 95–105 and 150 kDa MW. Reactivity to the 27, 50 and 75-95 kDa proteins were found more frequently in the IBS group compared to the HC’s (p < 0.001) and correlation was greater for Blastocystis-positive IBS patients (p < 0.001) than for negative IBS patients (p < 0.05). MAb1D5 reacted with proteins of 27 and 100 kDa, and anti-MMP-9 with 27, 50 and 75-100 kDa proteins. Bands were seen in zymograms around 100 kDa. Conclusions Low serum IgA levels are associated with Blastocystis carriage. All IBS patients were more likely to demonstrate reactivity with Blastocystis proteins of 27 kDa (likely a cysteine protease), 50 and 75-95 kDa MW compared to HC. The presence of antibodies to these Blastocystis proteins in some Blastocystis-negative subjects suggests either prior exposure to Blastocystis organisms or antibody cross reactivities. The anti-proMMP-9 reaction at 50 and 75–100 kDa and the zymogram result suggest that metalloproteases may be important Blastocystis antigens. Trial registration Australian and New Zealand Clinical Trials registry ACTRN: 12611000918921 Electronic supplementary material The online version of this article (doi:10.1186/s13071-015-1069-x) contains supplementary material, which is available to authorized users.
Collapse
Affiliation(s)
- Robyn Nagel
- School of Veterinary Science, The University of Queensland, Gatton Campus, Gatton, QLD, 4343, Australia. .,Toowoomba Gastroenterology Clinic, Suite 105 Medici Medical Centre, 15 Scott St, Toowoomba, QLD, 4350, Australia.
| | - Rebecca J Traub
- School of Veterinary Science, The University of Queensland, Gatton Campus, Gatton, QLD, 4343, Australia. .,Faculty of Veterinary and Agricultural Sciences, University of Melbourne, Parkville, VIC, 3052, Australia.
| | - Marcella M S Kwan
- Rural Clinical School, School of Medicine, The University of Queensland, Toowoomba, 4350, Australia.
| | - Helle Bielefeldt-Ohmann
- School of Veterinary Science, The University of Queensland, Gatton Campus, Gatton, QLD, 4343, Australia. .,Australian Infectious Diseases Research Centre, The University of Queensland, St. Lucia, QLD, 4072, Australia.
| |
Collapse
|
|
2
|
Blastocystis spp., Cryptosporidium spp., and Entamoeba histolytica exhibit similar symptomatic and epidemiological patterns in healthcare-seeking patients in Karachi. Parasitol Res 2012; 111:1357-68. [PMID: 22763702 DOI: 10.1007/s00436-012-2972-0] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2011] [Accepted: 05/15/2012] [Indexed: 12/14/2022]
Abstract
In this study, we collected data on the incidence of enteric parasites in healthcare-seeking individuals along with their symptoms to quantify the potential roles of factors such as age, sex, and seasonality in infection. We performed analysis to identify factors which could help differentiate parasitic infection from other causes of gastrointestinal illness in a community. The size of the patient population (n = 339), patient selection methodology, collection methods, and statistical analysis followed approaches from similar studies in core clinical journals. Ethical approval was obtained from the University of Karachi's Ethical Review Board. Fecal specimens (n = 339) submitted by symptomatic patients were collected from two clinical laboratories, along with information about the patients' age, sex, and symptoms. We found that symptoms of fever, vomiting, and constipation were 100 % predictive of finding a parasitic infection, while diarrhea was 88 % predictive of a parasitic infection. Gastrointestinal parasite-positive patients reported diarrhea (~60 %), vomiting (~30 %), fever (~25 %) and constipation (~25 %), while parasite-negative patients exhibited a symptomatic profile without fever, vomiting, and constipation. The distribution of symptoms in parasite-positive patients remained relatively invariant regardless of the parasite identified. Blastocystis spp.-mono-infected patients reported a similar profile to patients positive for Entamoeba histolytica/Entamoeba dispar and Cryptosporidium spp. Most parasitic infections exhibited a strong seasonal pattern, with a peak incidence in summer months. Infection by Blastocystis spp. was the most prevalent, and it was the only infection mathematically correlated to rainfall by Pearson's method. We observed no increase in healthcare-seeking behavior following a stressful community event, namely, the attempted assassination of Benazir Bhutto in Karachi. The data suggest that parasitological testing would produce a high yield of positive results when performed on healthcare-seeking patients in Karachi in 2007 with symptoms of fever, vomiting, or constipation and a low yield when performed on patients noting only abdominal pain. Parasitological testing also produces a higher yield on patients seen in summer months.
Collapse
|
|
3
|
The potential use of 29 kDa protein as a marker of pathogenicity and diagnosis of symptomatic infections with Blastocystis hominis. Parasitol Res 2010; 108:1139-46. [DOI: 10.1007/s00436-010-2156-8] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2010] [Accepted: 11/03/2010] [Indexed: 11/25/2022]
|
|
4
|
Xie X, McLean MD, Hall JC. Antibody-dependent cell-mediated cytotoxicity- and complement-dependent cytotoxicity-independent bactericidal activity of an IgG against Pseudomonas aeruginosa O6ad. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2010; 184:3725-33. [PMID: 20190144 DOI: 10.4049/jimmunol.0902732] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
In addition to Ag recognition, some Abs are capable of killing target organisms in the absence of phagocytes and complement. In this study, we report that an anti-Pseudomonas aeruginosa O6ad LPS IgG(1), tobacco-expressed human S20 IgG(1) (te-hS20), as well as its recombinant Fab and single-chain variable fragment (scFv) fragments have cellular- and complement-independent bactericidal activity. te-hS20 and its Fab and scFv significantly reduced viability of P. aeruginosa O6ad in dose- and time-dependent manners in vitro and also showed lower levels of bactericidal activity against P. aeruginosa PAO1, but had no activity against P. aeruginosa O10, Escherichia coli TG1, and Streptococcus agalactiae. The H chain and its Fd fragment both had significant Ag-binding and bactericidal activities against P. aeruginosa O6ad. Bactericidal activity was completely inhibited with specific LPS Ag, suggesting that Ag binding is involved in the bactericidal mechanism. Live/dead cell staining and electron microscopic observations indicate that the bactericidal effect was due to disruption of the cell wall and suggest inhibition of cell division. In addition to te-hS20, the Fab and scFv were also protective in vivo, as leukopenic mice had prolonged and improved survival after administration of these Ab fragments followed by challenge with P. aeruginosa O6ad cells at 80-90% lethal dose, supporting a bactericidal mechanism independent of phagocytes and complement. Understanding of the bactericidal mechanism will allow assessment of the potential for therapeutic application of these Abs.
Collapse
Affiliation(s)
- Xuemei Xie
- School of Environmental Sciences, University of Guelph, Guelph, Ontario, Canada
| | | | | |
Collapse
|
|
5
|
Wu B, Yin J, Texier C, Roussel M, Tan KSW. Blastocystis legumain is localized on the cell surface, and specific inhibition of its activity implicates a pro-survival role for the enzyme. J Biol Chem 2009; 285:1790-8. [PMID: 19915007 DOI: 10.1074/jbc.m109.049064] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023] Open
Abstract
Programmed cell death (PCD) is crucial for cellular growth and development in multicellular organisms. Although distinct PCD features have been described for unicellular eukaryotes, homology searches have failed to reveal clear PCD-related orthologues among these organisms. Our previous studies revealed that a surface-reactive monoclonal antibody (mAb) 1D5 could induce multiple PCD pathways in the protozoan Blastocystis. In this study, we identified, by two-dimensional gel electrophoresis and mass spectrometry, the target of mAb 1D5 as a surface-localized legumain, an asparagine endopeptidase that is usually found in lysosomal/acidic compartments of other organisms. Recombinant Blastocystis legumain displayed biphasic pH optima in substrate assays, with peaks at pH 4 and 7.5. Activity of Blastocystis legumain was greatly inhibited by the legumain-specific inhibitor carbobenzyloxy-Ala-Ala-AAsn-epoxycarboxylate ethyl ester (APE-RR) (where AAsn is aza-asparagine) and moderately inhibited by mAb 1D5, cystatin, and caspase-1 inhibitor. Interestingly, inhibition of legumain activity induced PCD in Blastocystis, observed by increased externalization of phosphatidylserine residues and in situ DNA fragmentation. In contrast to plants, in which legumains have been shown to play a pro-death role, legumain appears to display a pro-survival role in Blastocystis.
Collapse
Affiliation(s)
- Binhui Wu
- Department of Microbiology, Yong Loo Lin School of Medicine, Life Sciences Institute, National University of Singapore, Singapore 117597
| | | | | | | | | |
Collapse
|
|
6
|
Boorom KF, Smith H, Nimri L, Viscogliosi E, Spanakos G, Parkar U, Li LH, Zhou XN, Ok UZ, Leelayoova S, Jones MS. Oh my aching gut: irritable bowel syndrome, Blastocystis, and asymptomatic infection. Parasit Vectors 2008; 1:40. [PMID: 18937874 PMCID: PMC2627840 DOI: 10.1186/1756-3305-1-40] [Citation(s) in RCA: 120] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2008] [Accepted: 10/21/2008] [Indexed: 12/14/2022] Open
Abstract
Blastocystis is a prevalent enteric protozoan that infects a variety of vertebrates. Infection with Blastocystis in humans has been associated with abdominal pain, diarrhea, constipation, fatigue, skin rash, and other symptoms. Researchers using different methods and examining different patient groups have reported asymptomatic infection, acute symptomatic infection, and chronic symptomatic infection. The variation in accounts has lead to disagreements concerning the role of Blastocystis in human disease, and the importance of treating it. A better understanding of the number of species of Blastocystis that can infect humans, along with realization of the limitations of the existing clinical laboratory diagnostic techniques may account for much of the disagreement. The possibility that disagreement was caused by the emergence of particular pathogenic variants of Blastocystis is discussed, along with the potential role of Blastocystis infection in irritable bowel syndrome (IBS). Findings are discussed concerning the role of protease-activated receptor-2 in enteric disease which may account for the presence of abdominal pain and diffuse symptoms in Blastocystis infection, even in the absence of fever and endoscopic findings. The availability of better diagnostic techniques and treatments for Blastocystis infection may be of value in understanding chronic gastrointestinal illness of unknown etiology.
Collapse
Affiliation(s)
- Kenneth F Boorom
- Blastocystis Research Foundation, 5060 SW Philomath Blvd, #202, Corvallis, OR 97333, USA.
| | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
7
|
Tan KSW. New insights on classification, identification, and clinical relevance of Blastocystis spp. Clin Microbiol Rev 2008; 21:639-65. [PMID: 18854485 PMCID: PMC2570156 DOI: 10.1128/cmr.00022-08] [Citation(s) in RCA: 459] [Impact Index Per Article: 27.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
SUMMARY Blastocystis is an unusual enteric protozoan parasite of humans and many animals. It has a worldwide distribution and is often the most commonly isolated organism in parasitological surveys. The parasite has been described since the early 1900s, but only in the last decade or so have there been significant advances in our understanding of Blastocystis biology. However, the pleomorphic nature of the parasite and the lack of standardization in techniques have led to confusion and, in some cases, misinterpretation of data. This has hindered laboratory diagnosis and efforts to understand its mode of reproduction, life cycle, prevalence, and pathogenesis. Accumulating epidemiological, in vivo, and in vitro data strongly suggest that Blastocystis is a pathogen. Many genotypes exist in nature, and recent observations indicate that humans are, in reality, hosts to numerous zoonotic genotypes. Such genetic diversity has led to a suggestion that previously conflicting observations on the pathogenesis of Blastocystis are due to pathogenic and nonpathogenic genotypes. Recent epidemiological, animal infection, and in vitro host-Blastocystis interaction studies suggest that this may indeed be the case. This review focuses on such recent advances and also provides updates on laboratory and clinical aspects of Blastocystis spp.
Collapse
Affiliation(s)
- Kevin S W Tan
- Department of Microbiology, Laboratory of Molecular and Cellular Parasitology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Republic of Singapore.
| |
Collapse
|
|
8
|
Abstract
Among the waterborne protozoan parasites of medical and veterinary importance, Blastocystis is perhaps one of the less well-understood. However, in recent years, there has been a surge of interest in the organism, fueled in part by the possible association of Blastocystis infection with intestinal disorders, and its unusual taxonomic affiliations. Although there is information on the parasite's morphology, taxonomy and mode of transmission, its pathogenicity, life cycle, and function of certain organelles continue to baffle investigators. The clinical relevance of Blastocystis will be better answered once an animal model is found. Blastocystis infections have a worldwide distribution but prevalence is highest in areas with poor hygiene and deficient sanitation services and facilities. Application of modern molecular tools has advanced knowledge of the organism's genetic diversity, taxonomy and zoonotic potential.
Collapse
Affiliation(s)
- Kevin S W Tan
- Laboratory of Molecular and Cellular Parasitology, Department of Microbiology, Faculty of Medicine, National University of Singapore, 5 Science Drive 2, S 117597 Singapore, Singapore.
| |
Collapse
|
|
9
|
Wang KX, Li CP, Wang J, Cui YB. Epidemiological survey of Blastocystis hominis in Huainan City, Anhui Province, China. World J Gastroenterol 2002; 8:928-32. [PMID: 12378644 PMCID: PMC4656589 DOI: 10.3748/wjg.v8.i5.928] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To provide scientific evidence for prevention and controlling of blastocystosis, the infection of Blastocystis homonis and to study its clinical significance in Huainan City, Anhui Province, China.
METHODS: Blastocystis homonis in fresh stools taken from 100 infants, 100 pupils, 100 middle school students and 403 patients with diarrhea was smeared and detected with method of iodine staining and hematoxylin staining. After preliminary direct microscopy, the shape and size of Blastocystis homonis were observed with high power lens. The cellular immune function of the patients with blastocystosis was detected with biotin-streptavidin (BSA).
RESULTS: The positive rates of Blastocystis homonis in fresh stools taken from the infants, pupils, middle school students and the patients with diarrhea, were 1.0% (1/100), 1.0% (1/100), 0% (0/100) and 5.96% (24/403) respectively. Furthermore, the positive rates of Blastocystis homonis in the stool samples taken from the patients with mild diarrhea, intermediate diarrhea, severe diarrhea and obstinate diarrhea were 6.03% (14/232), 2.25% (2/89), 0% (0/17) and 12.31% (8/65) respectively. The positive rates of Blastocystis homonis in fresh stools of male and female patients with diarrhea were 7.52% (17/226) and 3.95% (7/177) respectively, and those of patients in urban and rural areas were 4.56% (11/241) and 8.02% (13/162) respectively. There was no significant difference between them (P > 0.05). The positive rates of CD3+, CD4+, CD8+ in serum of Blastocystis homonis-positive and-negative individuals were 0.64 ± 0.06, 0.44 ± 0.06, 0.28 ± 0.04 and 0.60 ± 0.05, 0.40 ± 0.05 and 0.30 ± 0.05 respectively, and the ratio of CD4+/CD8+ of the two groups were 1.53 ± 0.34 and 1.27 ± 0.22. There was significant difference between the two groups (P < 0.05, P < 0.01).
CONCLUSION: The prevalence of Blastocystis hominis as an enteric pathogen in human seems not to be associated with gender and living environment, and that Blastocystis hominis is more common in stool samples of the patients with diarrhea, especially with chronic diarrhea or obstinate diarrhea. When patients with diarrhea infected by Blastocystis hominis, their cellular immune function decreases, which make it more difficult to be cured.
Collapse
Affiliation(s)
- Ke-Xia Wang
- Department of Etiology and Immunology, School of Medicine, Anhui University of Science Technology, Huainan 232001, Anhui Province, China
| | | | | | | |
Collapse
|
|
10
|
Tan KSW, Singh M, Yap EH. Recent advances in Blastocystis hominis research: hot spots in terra incognita. Int J Parasitol 2002; 32:789-804. [PMID: 12062550 DOI: 10.1016/s0020-7519(02)00005-x] [Citation(s) in RCA: 90] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Despite being discovered more than 80 years ago, progress in Blastocystis research has been gradual and challenging, due to the small number of laboratories currently working on this protozoan parasite. To date, the morphology of Blastocystis hominis has been extensively studied by light and electron microscopy but all other aspects of its biology remain little explored areas. However, the availability of numerous and varied molecular tools and their application to the study of Blastocystis has brought us closer to understanding its biology. The purpose of this review is to describe and discuss recent advances in B. hominis research, with particular focus on new, and sometimes controversial, information that has shed light on its genetic heterogeneity, taxonomic links, mode of transmission, in vitro culture and pathogenesis. We also discuss recent observations that B. hominis has the capacity to undergo programmed cell death; a phenomenon similarly reported for many other unicellular organisms. There are still many gaps in our knowledge of this parasite. Although there is a growing body of evidence suggesting that B. hominis can be pathogenic under specific conditions, there are also other studies that indicated otherwise. Indeed, more studies are warranted before this controversial issue can be resolved. There is an urgent need for the identification and/or development of an animal model so that questions on its pathogenesis can be better answered. Another area that requires attention is the development of methods for the transfection of foreign/altered genes into B. hominis in order to facilitate genetic experiments.
Collapse
Affiliation(s)
- Kevin S W Tan
- Department of Microbiology, Faculty of Medicine, National University of Singapore, 5 Science Drive 2, Singapore 117597.
| | | | | |
Collapse
|