Intraoligochaete development of Myxobolus intimus (Myxosporea: Myxobolidae), a gill myxosporean of the roach (Rutilus rutilus)

The infection with Myxobolus intimus Zaika, 1965 in the gills of the roach Rutilus rutilus (L.) from Lake Balaton was recorded in 28 out of the 39 fish examined. Developing and mature plasmodia were detected on the gills exclusively in the spring. The Myxobolus intimus infection was found only in 2- to 3-year-old fish. In histological sections, young plasmodia were found in capillaries of the secondary lamellae. More mature, round plasmodia 0.4-0.6 mm in diameter, deformed the respiratory lamellae. The intraoligochacte development of M. intimus was studied in experimentally infected oligochaetes. In two experiments, uninfected Tubifex tubifex Muller and Limnodrilus hoffmeisteri (Claparede) were exposed to mature myxospores of M. intimus. In both experiments, typical triactinospores developed in T. tubifex specimens but no infection was found in L. hoffmeisteri. In semithin sections, developmental stages, pansporocysts and actinospores, were found within the proliferated gut epithelium of T. tubifex. Triactinospores were first released from oligochaetes 37 and 58 days after initial exposure in the two experiments, respectively. Each triactinospore had three pyriform polar capsules and a cylindrical sporoplasm with 32 secondary cells. The spore body joined the 3 caudal projections with a moderately long style

Radiodiagnostic method for studying the dynamics of Anguillicola crassus (Nematoda: Dracunculoidea) infection and pathological status of the swimbladder in Lake Balaton eels

Swimbladder changes resulting from Anguillicola crassus infection of the European eel Anguilla anguilla have been the subject of several studies reported in the literature. These investigations, however, studied exclusively the status of infection at a given point in time and did not deal with changes in swimbladder infection in eels suffering from anguillicolosis over a period of time. In this study, A. crassus-induced pathological changes were monitored in 78 eels naturally infected in Lake Balaton and subsequently kept in the laboratory, thus excluding the possibility of further infection. During the 3 mo study, the status of the swimbladder was checked by radiographic examination on 4 occasions. At the end of the study the eels were dissected and the gross pathological changes in the swimbladders were compared with the radiographic findings. As compared to their starting condition, by the end of the study the pathological status of the swimbladder had deteriorated in 55 % and remained the same in 37 % of the cases. Tendency to improvement (1 %) and variable findings (7 %) were recorded in a low percentage of cases only. With the help of the radiographs presented, the dynamics of A. crassus infection and of changes in the swimbladder of individual eel specimens can be monitored easily

A Balaton és vízrendszere halfajainak parazitológiai vizsgálata = Parasitological investigations on fishes of Lake Balaton and its tributaries

A kutatás négy éve alatt a Balaton és vízrendszere több mintavételi helyén rendszeres halbefogásokkal követtük nyomon a halak parazita-fertőzöttségét. Parazita-anyagot (elsősorban nyálkaspórás és coccidium-fajokat) gyűjtöttünk kísérletes- és molekuláris biológiai vizsgálatokhoz, az egyes parazita-fajok szokásos évszakos megjelenésének idejében évente ismétlődően, melyek egy részét laboratóriumi kísérleteinkben használtunk fel. Egy nyálkaspórás faj intraoligochaeta-fejlődésmenetét tisztáztuk, valamint bizonyítottuk a copepoda-rákok actinosporák eliminálásában való szerepét. Kísérletesen vizsgáltuk, hogy az anguillicolózis hogyan befolyásolja az angolnák vándorló-képességét, ill. vizsgáltuk az úszás hatását az angolna petefejlődésére. Röntgendiagnosztikai eljárást alkalmaztunk az egyes angolna egyedek úszóhólyag-fertőzöttsége dinamikájának, ill. az úszóhólyagok regenerációs képességének megállapítása céljából. Vizsgáltuk a vérélősködő métely fajok halakban való fejlődését és kórtani hatását. Adatokat gyűjtöttünk a kokcidium fajokkal való fertőződés dinamikájára és kísérletesen tanulmányoztuk, hogy az oligochaeták mellett milyen vízi szervezetek játszhatnak szerepet a gócos kokcidiózis évszakos megjelenésében. Új eredményeink többségét tudományos lapokban jelentettük meg. Az OTKA téma részleges segítségével több külföldi kooperációban készült munka is megvalósult (Portugál, Maláj, Szír-együttműködések) | During the four-year research we monitored the parasitic infections of fishes by regular fish samplings of Lake Balaton and its tributaries. We collected parasite material (primarily myxosporeans and coccidians) for experimental and molecular studies annually, at the time of the usual seasonal appearance of the different parasite species. We have clarified the intraoligochaete stage of development of a myxosporean species of roach and demonstrated the role of copepods in the elimination of actinospores. We conducted experiments to study the impact of anguillicolosis on the migrating ability of eels and the effect of swimming on ovarian development in eels. We applied a radiodiagnostic technique to determine the dynamics of swimbladder infection in individual eel specimens and the regenerating ability of the swimbladder. We also studied the intrapiscine development and pathological effect of blood-parasitic fluke (Sanguinicola) species. We collected data on the dynamics of infection with coccidium species and conducted experiments to determine which aquatic organisms, other than oligochaetes, may play a role in the seasonal occurrence of nodular coccidiosis. The majority of our new results have been published in scientific journals. Several co-operative projects with foreign researchers (Portuguese, Malaysian and Syrian co-operations) have also been implemented, with partial financial support from the National Scientific Research Fund of Hungary (OTKA)

A halélősködő nyálkaspórások fejlődésének, kórtanának és fajlagosságának vizsgálata kísérletes és molekuláris biológiai módszerekkel = Studies on the development, pathogenicity and host specificity of fish myxosporeans by experimental and molecular biological methods

Balatonból, Kis-Balatonból, Dunából és Tiszából, valamint néhány tógazdaságból gyűjtött halon vizsgáltuk a nyálkaspórás fertőzöttség alakulását, és ugyanezen biotópokon tanulmányoztuk a nyálkaspórások oligochaeta férgekben élő actinospóra stádiumainak előfordulását. Magyarországon és külföldön gyűjtött mintáink alapján több új Myxosporea fajt írtunk le, illetve jellemeztük a kevésbé ismert fajok myxospóráinak morfológiai tulajdonságait. Hasonló vizsgálatokat végeztünk a nyálkaspórások oligochaeta férgekben fejlődő actinospóra stádiumain, illetve a férgekből kirajzott actinospórákon is. Szövettani és molekuláris módszerekkel vizsgáltuk néhány Myxosporea szerv- ill. szövetspecificitását, és a domolykó fertőzöttségét modellként használva 8 faj esetében részleteztük a különböző lokációkban fejlődő fajok morfológiai és molekuláris különbségét. Néhány Myxobolus faj esetében laboratóriumi kísérletekben reprodukáltuk a teljes fejlődési ciklust, azonban ez csak a M. intimus fajjal volt több alkalommal megismételhető. A nehezen kivitelezhető, hosszadalmas és bizonytalan kísérletes munkát fokozatosan molekuláris technikákkal helyettesítettük, és a talált, morfológiailag jellemzett myxospóra és actinospóra alakok 18S rDNS szerkezetének tanulmányozásával választottuk ki a megfelelő actinospóra-myxospóra párokat. Kísérletesen igazoltuk, hogy a copepoda rákok a lebegő actinospórákat a vízből kiszűrik. A világon eddig leírt 751 faj alapján elkészítettük Myxobolus genus synopsisát. | Myxosporean infection of fishes was examined in Danube and Tisza rivers, in Lake Balaton and Kis-Balaton and in some fish farms. In the same biotopes the occurrence of actinosporean stages of myxosporeans in oligochaetes was also studied. Based on samples collected in Hungary and abroad some new myxosporean spp. were described and some less known species were morphologically characterised. Similar morphological studies were performed on actinosporean stages of myxosporeans developing in oligochaetes and on floating actinospores emerged from the worms. Organ, site and tissue specificity of several myxosporean spp. was studied by histological and molecular genetic methods, and using myxosporean infection of the chub (Leuciscus cephalus) as a model, morphological and genetical differences were characterised for 8 species developing in different locations in this fish. Trials were performed for reproducing the complete developmental cycle of selected Myxobolus spp. and in case of M. intimus these experiment were several times successfully repeated. The tiresome life cycle experiments were step by step substituted by molecular techniques and the adequate myxospore/actinospore pairs were identified by studying their 18S rDNA sequences. It was experimentally proved that copepods can filter floating actinospores from the water. We have participated in preparing a synopsis on the 751 Myxobolus species described in the world up to this time

Rapid detection of Listeria monocytogenes in raw milk and soft cheese by a redox potential measurement based method combined with real-time PCR

The incidence of outbreaks of foodborne listeriosis has indicated the need for a reliable and rapid detection of the microbe in different foodstuffs. A method combining redox potential measurement and real-time polymerase chain reaction (PCR) was developed to detect Listeria monocytogenes in artificially contaminated raw milk and soft cheese. Food samples of 25 g or 25 ml were homogenised in 225 ml of Listeria Enrichment Broth (LEB) with Oxford supplement, and the redox potential measurement technique was applied. For Listeria species the measuring time was maximum 34 h. The absence of L. monocytogenes could reliably be proven by the redox potential measurement method, but Listeria innocua and Bacillus subtilis could not be differentiated from L. monocytogenes on the basis of the redox curves. The presence of L. monocytogenes had to be confirmed by real-time PCR. The combination of these two methods proved to detect < 10 cfu/g of L. monocytogenes in a cost- and time-effective manner. This method can potentially be used as an alternative to the standard nutrient method for the rapid detection of L. monocytogenes in food

Fluorescent probes for the dual investigation of MRP2 and OATP1B1 function and drug interactions

Detoxification in hepatocytes is a strictly controlled process, in which the governed action of membrane transporters involved in the uptake and efflux of potentially dangerous molecules has a crucial role. Major transporters of hepatic clearance belong to the ABC (ATP Binding Cassette) and Solute Carrier (SLC) protein families. Organic anion-transporting polypeptide OATP1B1 (encoded by the SLCO1B1 gene) is exclusively expressed in the sinusoidal membrane of hepatocytes, where it mediates the cellular uptake of bile acids, bilirubin, and also that of various drugs. The removal of toxic molecules from hepatocytes to the bile is accomplished by several ABC transporters, including P-glycoprotein (ABCB1), MRP2 (ABCC2) and BCRP (ABCG2). Owing to their pharmacological relevance, monitoring drug interaction with OATP1B1/3 and ABC proteins is recommended. Our aim was to assess the interaction of recently identified fluorescent OATP substrates (various dyes used in cell viability assays, pyranine, Cascade Blue hydrazide (CB) and sulforhodamine 101 (SR101)) (Bakos et al., 2019; Patik et al., 2018) with MRP2 and ABCG2 in order to find fluorescent probes for the simultaneous characterization of both uptake and efflux processes. Transport by MRP2 and ABCG2 was investigated in inside-out membrane vesicles (IOVs) allowing a fast screen of the transport of membrane impermeable substrates by efflux transporters. Next, transcellular transport of shared OATP and ABC transporter substrate dyes was evaluated in MDCKII cells co-expressing OATP1B1 and MRP2 or ABCG2. Our results indicate that pyranine is a general substrate of OATP1B1, OATP1B3 and OATP2B1, and we find that the dye Live/Dead Violet and CB are good tools to investigate ABCG2 function in IOVs. Besides their suitability for MRP2 functional tests in the IOV setup, pyranine, CB and SR101 are the first dual probes that can be used to simultaneously measure OATP1B1 and MRP2 function in polarized cells by a fluorescent method. © 2020 The Author(s