Oh my aching gut: irritable bowel syndrome, Blastocystis, and asymptomatic infection

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

Project on the stimulation of human mononuclear cells by prothymosin a

In the framework of this thesis, the effect of prothymosin a on the proliferation and cytotoxic functions of anti-CD3 stimulated human peripheral blood mononuclear cells (PBMC) and its utility as a stimulating agent in adoptive immunotherapy protocols was investigated. The mechanisms by which prothymosin a mediates its immunomodulatory effects, were also explored. Anti-CD3 stimulated PBMC from healthy individuals and cancer patients showed increased proliferative responses and cytotoxic ability against tumor cell lines when incubated with prothymosin a. The effect was more prominent in the case of PBMC isolated from cancer patients. Prothymosin a also potentiated cytotoxicity against autologous cancer cells, but not against autologous blast cells. This phenomenon was non-MHC restricted and was mediated through CD8+ and CD56+ lymphocytes. Secretion of IL- lß, IL-2 and TNF-a, (but not IL-7 and GM-CSF) was significantly increased in the presence of prothymosin a; none of these cytokines however is solely responsible for the augmented cytotoxicity. CD2, CD 18, CD54, CD49f and perforin expression was also enhanced and a direct involvement of these antigens in the cytotoxic process induced by prothymosin a was demonstrated. The ratio of CD8, CD4, and CD56 subpopulations did not change while surface expression of CD25 was up-regulated in all types of cells in the presence of prothymosin a, which thereby exhibited a stimulatory effect on CD4+ T-lymphocytes as well. Furthermore, a neutral effect on the balance between Type1 and Type2 immune responses was observed. In conclusion, data in the present thesis strongly substantiate the role of prothymosin a as an immunomodulatory agent and promote its prospective use in clinical trials for cancer immunotherapy.Στα πλαίσια της παρούσης διατριβής εξετάστηκε η επίδραση της προθυμοσίνης α σε ενεργοποιημένα με αντί-CD3 μονοκλωνικό αντίσωμα ανθρώπινα μονοπύρηνα κύτταρα περιφερικού αίματος και η δυνατότητα χρησιμοποίησης της σε πρωτόκολλα ανοσοθεραπείας σε ασθενείς με καρκίνο. Επίσης εξετάσθηκαν οι μηχανισμοί μέσω των οποίων ασκεί την ανοσορυθμιστική δράση. Η προθυμοσίνη α βρέθηκε να αυξάνει τον ρυθμό πολλαπλασιασμού και την κυτταροτοξική δράση των ενεργοποιημένων με αντί-CD3 μονοκλωνικό αντίσωμα ανθρώπινων λεμφοκυττάρων προερχόμενα από υγιείς δότες και ασθενείς με καρκίνο. Το ανοσοενισχυτικό αποτέλεσμα της προθυμοσίνης α είναι εντονότερο σε λεμφοκύτταρα προερχόμενα από καρκινοπαθείς, τα οποία εμφανίζουν χαμηλότερο βαθμό ενεργοποίησης από τα λεμφοκύτταρα των υγιών. Επιπλέον αυξάνεται η κυτταροτοξικότητα εναντίον αυτόλογων καρκινικών κυττάρων και όχι εναντίον βλαστικών κυττάρων. Η αυξημένη κυτταροτοξική δράση που επάγεται από την προθυμοσίνη α, είναι μη εξαρτώμενη από μόρια του Μείζονος Συμπλέγματος Ιστοσυμβατότητας και μεσολαβείται από CD8+ και CD56+ κύτταρα. Στο υπερκείμενο κυτταροκαλλιεργειών με προθυμοσίνη α ήταν σημαντικά αυξημένη η συγκέντρωση των κυτταροκινών IL-1 β, IL-2 και TNF-α. Η δράση της προθυμοσίνης α όμως δεν υποκαθίσταται από τις συγκεκριμένες κυτταροκίνες καθώς καμία από μόνη της, δεν αυξάνει την κυτταροτοξική δράση των λεμφοκυττάρων στον ίδιο βαθμό. Τα λεμφοκύτταρα παρουσίασαν αυξημένη έκφραση των CD2, CD18, CD54, CD49f και της ενδοκυτταρικής περφορίνης, και σχετίζονται άμεσα με την αυξημένη κυτταροτοξική δράση των λεμφοκυττάρων. Η έκφραση του CD25 αυξάνεται σε όλους τους υποπληθυσμούς των Τ-λεμφοκυττάρων και η αναλογία των CD4+, CD8+, και CD56+ κυττάρων δεν μεταβάλλεται ενδεικτικό ανοσοενισχυτικής δράσης της προθυμοσίνης α και στα CD4+ κύτταρα. Επιπλέον, δεν παρατηρείται μεταβολή της ισορροπίας Thl / Th2 καθώς οι συγκεντρώσεις των κυτταροκινών IL-6, IL-12, και IFN-γ δεν μεταβάλλονται παρουσία της προθυμοσίνης α. Η ικανότητα της προθυμοσίνης α να αυξάνει την κυτταροτοξική ικανότητα των λεμφοκυττάρων εναντίον ετερόλογων και αυτόλογων καρκινικών κυττάρων, χωρίς να μεταβάλλει το λειτουργικό τους πρόγραμμα, την καθιστά υποψήφιο μόριο για κλινικές δοκιμές στην ανοσοθεραπεία του καρκίνου

Occurrence of Cryptosporidium and Giardia in recycled waters used for irrigation and first description of Cryptosporidium parvum and C-muris in Greece

Here, we present the first time findings regarding the occurrence of Cryptosporidium and Giardia in sewage waters and the first molecular characterization of Cryptosporidium species in Greece. Biological treatment plants from three regions in Greece have been investigated. The detection of Cryptosporidium oocysts was by modified Ziehl-Neelsen acid fast (MZN-AF) and by immunofluorescence microscopy (IFT) for Cryptosporidium and Giardia (oo)cysts, whereas nested PCR based on the SSU rDNA assay was used for molecular detection of Cryptosporidium followed by sequencing for the genetic characterization of the species. In total, 73 samples (37 raw sewage samples and 38 of treated water samples) were collected and analyzed. Of the 73 water samples, 4 samples were Cryptosporidium-positive by IFT and staining, 12 samples were Cryptosporidium-positive by nested PCR; 9 samples were Giardia-positive by IFT. We showed that Cryptosporidium cysts are found both in the input and the discharge of the biological treatment plants. Molecular characterization of Cryptosporidium based on the small subunit ribosomal DNA gene resulted in the determination of Cryptosporidium parvum and Cryptosporidium muris Greek isolates. This is the first report of Cryptosporidium and Giardia occurrence in wastewaters and the first molecular identification of Cryptosporidium species in Greek environments. As the treated water is used for irrigation, or it is discharged into the sea, our findings indicate that biological treatment facilities constitute a possible risk for public health because the related species are prevalent in humans; the results invite for further epidemiological investigations to evaluate the real public health risk in Greece

Molecular screening of cat and dog ectoparasites for the presence of Bartonella spp. in Attica, Greece.

The purpose of this study was the molecular detection of Bartonella spp. in fleas and ticks parasitizing cats and dogs from 39 locations in Attica, Greece. One hundred and forty five ectoparasites (104 fleas and 41 ticks) from 92 cats and 53 dogs were investigated individually using PCRs targeting the 16S-23S ribosomal RNA intergenic spacer (ITS) and the citrate synthase (gltA) genetic loci. Bartonella spp. were detected in 14 out of 104 fleas (13.5%) and in none of the ticks examined. Consequent sequence analysis of the amplicons from the two loci identified 3 strains as Bartonella henselae, and 11 as Bartonella clarridgeiae. Οur study demonstrates the presence of B. henselae and B. clarridgeiae in Ctenocephalides felis fleas from cat and dog in Greece. We also report a novel ITS sequence for B. clarridgeiae. Considering that fleas could pose a risk for human bartonellosis from their infected hosts, further studies on the public health risk of Bartonella presence in animal ectoparasites are warranted

Visceral leishmaniasis and COVID-19 coinfection - A case report

As the COVID-19 pandemic spreads across the globe, it will undoubtedly cross paths with long endemic infectious diseases in different areas. Interactions between SARS-CoV2 and well-known pathogens will likely give rise to unfamiliar clinical presentations, depending on complex and as yet unknown immunological interactions. We present a case of coinfection with COVI19 and visceral leishmaniasis and discuss recent reports regarding coexistence of SARS-CoV2 and Leishmania spp. to date. (C) 2021 Published by Elsevier Ltd

Phlebotomine sandflies and factors associated with their abundance in the leishmaniasis endemic area of Attiki, Greece

Leishmaniasis is a parasitic disease of animals and humans caused by several Leishmania species and transmitted by phlebotomine sandflies. The aim of the present study was to identify the species of field collected phlebotomine sandflies in the endemic area of the Attiki during 4 consecutive years, to isolate the Leishmania parasites from the infected sandflies, and identify possible factors associated with sandfly abundance in the area. A total of 542 trappings were made in 46 collection sites, in purely urban areas, periurban areas, and purely rural areas in Attiki. Out of the 3254 sandflies trapped, 1448 (44.43%) were female and 241 (16.64%) of the females were blood fed while Leishmania infantum DNA was detected in the 0.41% of them. Regarding sandfly species, the most prevalent was Phlebotomus tobbi (41.52%) followed by Sergentomyia minuta (27.44%), P. neglectus (14.83%), P. simici (11.08%), P. papatasi (3.68%), P. similis (0.89%), and P. alexandri (0.56%). Periurban areas were found to have the highest density of sandfly populations

Direct detection of Blastocystis sp in human faecal samples and subtype assignment using single strand conformational polymorphism and sequencing

Blastocystis is an anaerobic parasitic microorganism, which has been found in the intestinal tract of many vertebrates including humans. Recently, members of Blastocystis sp. were classified into nine subtypes, based on phylogenetic trees derived from sequence analysis of the small subunit ribosomal RNA (SSU rRNA) gene. The role of Blastocystis in human disease remains uncertain and the existence of pathogenic and non-pathogenic subtypes is under investigation. We report the development of a polymerase chain reaction (PCR)-based assay that is able to detect Blastocystis directly from human faeces. Furthermore, combined with single strand conformational polymorphism (SSCP) analysis and/or sequencing of the respective PCR product, the protocol can classify Blastocystis among the nine established subtypes. The method was applied to 45-positive and 30-negative faecal samples and proved to be highly sensitive and specific. Genotyping using SSCP analysis and sequencing revealed that subtype 3 is the most frequent in Greece, while subtypes 1, 2, 4, 6 and 7 are also present but in lower frequencies. Hopefully, the simplicity of the proposed method will contribute toward large-scale epidemiological studies for prompt clarification of the role of the parasite. (C) 2007 Elsevier Ltd. All rights reserved