Detection of Anaplasma dna in Ixodes ricinus ticks: pitfalls

A total of 150 nymphal Ixodes ricinus (L., 1758) (Acari: Ixodidae) from the Czech Republic were examined for Anaplasma phagocytophilum (Foggie, 1949) Dumler et al., 2001 by PCR using EHR521/747 primers: 22 of 50 pools were positive (minimum prevalence, 14.7%). However, sequencing of the PCR products did not show complete homology with A. phagocytophilum (91%) while the closest relationship (95%) was found to "Candidatus Ehrlichia walkefii". The results indicate a need for care in interpretation of Anaplasma PCR results and for PCR optimization for detecting A. phagocytophilum in ticks.     

Xenoma-like formations induced by Soricimyxum fegati (Myxosporea) in three species of shrews (Soricomorpha: Soricidae), including records of new hosts

In South Bohemia, Czech Republic, 178 shrews, including 98 common shrews, Sorex araneus L., 70 pygmy shrews, Sorex minutus L., and 10 lesser white-toothed shrews, Crocidura suaveolens (Pallas), were examined for Soricimyxum fegati Prunescu, Prunescu, Pucek et Lom, 2007 infections, using squash preparations of unfixed tissues, histological sections and molecular methods. The infection was found in 51 (52%) S. araneus, 14 (20%) S. minutus and 1 (10%) C. suaveolens. The records of the latter two species extend host range of S. fegati. Lesions associated with S. fegati infections in the liver, the organ of specific localisation of the parasite, were found to be induced by proliferative stages migrating toward lumina of bile ducts. In other organs of these three host species, xenoma-like formations (XLFs) were found that severely injured blood vessels. XLFs contained presporogonic stages of S. fegati, whose species identity was evidenced using molecular methods.     

Population genetic structure of Mycosphaerella graminicola and Quinone Outside Inhibitor (QoI) resistance in the Czech Republic

Damage caused by the wheat pathogen Mycosphaerella graminicola increased rapidly during the last two decades in the Czech Republic. We collected isolates from naturally infected fields in seven wheat-growing locations and analysed these using eight microsatellite markers. All markers were highly polymorphic. We found a high degree of genetic diversity and low clonality within all sampled Czech populations. We identified 158 unique multilocus haplotypes among 184 isolates. Field populations showed weak genetic structure but we detected more differentiation between climatic regions within the Czech Republic. We compared the Czech field populations to populations from the United Kingdom, Germany and Switzerland and found a marked differentiation between Czech populations and Western European populations. We hypothesize that decades of different agricultural practices, including the use of different wheat cultivars, may explain this genetic differentiation. We detected a rapid increase in QoI fungicide resistance during the sampling period from 2005 to 2011, coinciding with the widespread application of this class of fungicides in the Czech Republic. M. graminicola populations in the Czech Republic underwent a rapid adaptive evolution from sensitivity to resistance similar to what was described earlier in Western Europe.     

Bartonella infections in fleas (Siphonaptera: Pulicidae) and lack of bartonellae in ticks (Acari: Ixodidae) from Hungary

Fleas (95 Pulex irritans, 50 Ctenocephalides felis, 45 Ctenocephalides canis) and ixodid ticks (223 ixodes ricinus, 231 Dermacentor reticulatus, 204 Haemaphysalis concinna) were collected in Hungary and tested, in assays based on PCR, for Bartonella infection. Low percentages of P. irritans (4.2%) and C. felis (4.0%) were found to be infected. The groEL sequences of the four isolates from P. irritans were different from all the homologous sequences for bartonellae previously stored in GenBank but closest to those of Bartonella sp. SE-Bart-B (sharing 96% identities). The groEL sequences of the two isolates from C. felis were identical with those of the causative agents of cat scratch disease, Bartonella henselae and Bartonella clarridgeiae, respectively. The pap31 sequences of B. henselae amplified from Hungarian fleas were identical with that of Marseille strain. No Bartonella-specific amplification products were detected in C. canis, I. ricinus, D. reticulatus and H. concinna pools.     

Molecular characterization of <Emphasis Type="Italic">Xiphinema brevicollum</Emphasis> (Nematoda: Longidoridae) from the Czech Republic

Populations of Xiphinema brevicollum occurring in the Czech Republic were described morphologically and molecularly. Published species-specific primer set BL18 and BV3 was used to amplify three populations of X. brevicollum from the Czech Republic. These primers were tested against 9 species of Xiphinema and 11 species of Longidorus. Amplification was also observed for X. inaequale, X. italiae and X. lambertii. Three additional markers, mitochondrial cytochrome c oxidase subunit 1, ribosomal D2/D3 expansion segment of the 28S gene and 18S gene, were amplified and sequenced for X. brevicollum, X. inaequale and X. lambertii belonging to X. americanum-group. Comparison of cox1 sequences of X. brevicollum from the Czech Republic with X. taylori from the Slovak Republic (accession number AM086702) suggested that these populations represent the same species.     

Fatal experimental Babesia microti infections in the Norwegian lemming, Lemmus lemmus (L.).

Experimental infections of Babesia microti in laboratory-reared Clethrionomys glareolus revealed that approximately 15% of the erythrocytes were infected with single ring forms during peak parasitemia. Infected erythrocytes could be detected in blood smears up to one month post infection. C. glareolus treated with a single injection of Depo-Medrol i.m. two days prior to infection displayed a four-fold increase in number of infected erythrocytes at peak parasitemia, 35% of which contained more than one Babesia, and a prolongation of the infection. B. microti infections in 35 laboratory--reared Lemmus lemmus were fatal. Multiple invasion of erythrocytes, anemia, icterus, hemoglobinuria, anorexia and weight loss, and adrenal and splenic hypertrophy were characteristic for B. microti infections in Norwegian lemmings.     

Distribution and Ecology of the Russian Wheat Aphid,Diuraphis noxia (Kurdj.), expanded to Central Europe (Hom.: Aphididae)

A part of the territory of the Czech Republic represents the up-dated northern limit of distribution (status 1997) which the Russian wheat aphid (=RWA),Diuraphis noxia (Kurdj.) has reached during its expansion from the southeast to central Europe. The newly defined area is classified as one of the adventive routes which may be derived from the main expansion of RWA from its native home (central-western Asia) into the broader Mediterranean area. Distribution and its history, and ecology (life-cycle, host plants, seasonal history, both native and introduced natural enemies) of RWA are presented on ground of field evidence and trials obtained in 1995–1997 in the Czech Republic. The origin of RWA, its distribution and establishment in the target area, as well as prognosis of its further expansion, are reviewed and discussed. Steps and approaches to RWA detection in the crop, as well as some notes on its management, are added.     

Temporal and spatial distribution of glochidial larval stages of European unionid mussels (Mollusca: Unionidae) on host fishes

Glochidia are the larval stage of freshwater unionid mussels that parasitize the fins and gill apparatus of fish. A total of 22 fish species were examined for the presence of glochidia whose distribution on individual hosts was studied on three common fish species, the roach Rutilus rutilus (L.), perch Percafluviatilis L. and bitterling Rhodeus sericeus (Pallas). Between 1997 and 1999, the fish were obtained from the rivers Morava and Kyjovka and surrounding water pools in the Czech Republic. The glochidia of two genera, Unio and Anodonta, were found. Anodonta glochidia were observed on 10 fish species, Unio glochidia on 17 fish species. There was a difference in spatial distribution of glochidia on the body of the host fish. Unio glochidia were predominantly located on the gills, whereas most Anodonta glochidia were found on the fins, with the highest numbers of glochidia were observed on the margin of the pectoral fins. For the gill apparatus, Unio glochidia were found predominantly on the second and third arch. Anodonta glochidia were predominantly found during winter and spring (November-May), whereas Unio glochidia were more abundant during May and June. The number of glochidia was positively correlated with fish length in perch highly infected by Anodonta glochidia and perch infected by Unio glochidia. Of the three fish species, the highest occurrence of parasites was found on perch with fewer observed on roach. In spite of the close relationship between bitterling and unionid mussels, glochidiosis was rare on this fish species.     

Recent surveys for Bursaphelenchus xylophilus in the Czech Republic1

The State Phytosanitary Administration of the Czech Republic (SPA) conducted a survey of Bursaphelenchus xylophilus in 2004 and 2005. In 2004 97 locations were surveyed and 60 samples were taken. B. xylophilus was not found in any of the samples. In 2005, the SPA performed 120 phytosanitary inspections on solitary conifers, and in forest stands and clumps of trees. Checks were performed on Pinus sylvestris and Pinus nigra trees and in coniferous woodland mainly in areas at risk of invasion from B. xylophilus. Of 70 phytosanitary inspections performed in at risk areas, 13 samples were taken for laboratory processing and analysis. Fifty phytosanitary inspections were carried out outside risk areas, mainly in forests and parks, and 28 samples were taken. The Czech University of Agriculture in Prague together with the diagnostic laboratory of the SPA carried out a second survey in 2005. Fifty‐eight samples were taken from 58 surveyed locations in forests where coniferous trees with symptoms had been found. Laboratories performed a survey of B. xylophilus vectors to find only one vector species, Monochamus galloprovincialis subsp. pistor. In total in 2005, 178 phytosanitary inspections were carried out in the Czech Republic. B. xylophilus was not detected in either samples or vectors. Thus the results of the surveying were negative in both 2004 and 2005. The status of Bursaphelenchus xylophilus in the Czech Republic is: absent, confirmed by survey.     

Flight patterns of Russian wheat aphid,Diuraphis noxia (Kurdj.) during its expansion to Central Europe (Hom., Aphididae)

Flight patterns of migrant alate Russian wheat aphids (=RWA),Diuraphis noxia (Kurdj.) during its expansion from the south-east to Central Europe were determined. Samples were taken by Rothamsted-type suction air traps in Hungary (one location, 1990–1997) and in the Czech Republic (5 locations, 1994–1998). The results document a progressive expansion of RWA, and population variation in the individual years and periods of the season. A comparison of the evidence on RWA in the individual years since its detection in Hungary and in the Czech Republic supports the presumption on the occurrence of an expansion route from the south-east (Turkey) to Central Europe. This expansion is classified as an adventive route of the over-all expansion that has covered the whole West-Mediterranean (South Europe, North Africa).     

Variation and distribution of virulence phenotypes of Bremia lactucae in natural populations of Lactuca serriola

During 1997–2000 the virulence variation and distribution of virulence phenotypes of Bremia lactucae (lettuce downy mildew) in natural populations of Lactuca serriola (prickly lettuce) were studied. Altogether 139 isolates of B. lactucae originating from the Czech Republic, France and Germany were examined for the presence of 27 virulence factors (v-factors) and their combinations. In the Czech population, 37 different v-phenotypes (P1–P37) of B. lactucae were found to occur on L. serriola . Most v-phenotypes were characterized by v-factors that match resistance ( Dm genes/R-factors) carried by L. serriola . A wide diversity of v-phenotypes was recorded every year, but most were rare and did not reappear in other populations of B. lactucae . The three v-phenotypes P21, P28 and P29 were most frequent and widely distributed. There was variation both between spatially isolated populations and within populations of the pathogen. Geographic differences in virulence were found for the southern parts of Moravia, where the phenotypic composition of pathogen populations was completely different from the remaining part of the area investigated. However, some populations had v-phenotypes similar to those of spatially distant populations in Moravia.     

Fungicide sensitivity of populations of wheat powdery mildew (Erysiphe graminis f.sp. Tritici) in Central Europe in 1993

In 1993 we observed the sensitivity of wheat powdery mildew populations (Erysiphe graminis DC f.sp. tritici Marchal) from the Czech Republic, Austria, Hungary and Slovakia to the fungicides triadimenol, tebuconazole, propiconazole, flutriafol and fenpropimorph. The highest resistance value was shown to triadimenol, which attained a mean resistance factor (MRF) of 29 (expressing how many times the population is more resistant than are standard sensitive isolates) in the mildew population from the Czech Republic. The mildew populations from eastern Slovakia and eastern Hungary, populations geographically isolated from the other populations, showed very high sensitivity to all fungicides tested. There was most sensitivity to fenpropimorph (smallest MRF values) compared with the other fungicides. Cross-resistance was established among all triazoles used, but not between triazoles and fenpropimorph. Sensitivity of wheat powdery mildew populations from Central Europe to these fungicides is considered adequate, and the development of resistance has shown a decreasing tendency in recent years.     

Hymenopterous parasitoids of the cherry bark tortrix, Enarmonia formosana (Scopoli) in central-east Europe (Hymenoptera, Ichneumonoidea; Lepidoptera, Tortricidae)

Strategies targeting the biological control of the cherry bark tortrix (=CBT), Enarmonia formosana (Scopoli) in North America have also included the search for potentially useful native hymenopterous parasitoids of the pest in Europe. The search for such agents undertaken in the central-east Europe (Czech Republic, Slovak Republic, Hungary) resulted in determination of several ichneumonid species (Campoplex cf. dubitator Horst., Liotryphon sp., Lissonotus sp., Pimpla spuria (Grav.), Pimpla turionellae (L.), and some others, and a braconid (Apanteles britannicus Wilk.) in 2001–2002.     

Hymenopterous parasitoids of the cherry bark tortrix, Enarmonia formosana (Scopoli) in central-east Europe (Hymenoptera, Ichneumonoidea; Lepidoptera, Tortricidae)

Strategies targeting the biological control of the cherry bark tortrix (=CBT), Enarmonia formosana (Scopoli) in North America have also included the search for potentially useful native hymenopterous parasitoids of the pest in Europe. The search for such agents undertaken in the central-east Europe (Czech Republic, Slovak Republic, Hungary) resulted in determination of several ichneumonid species (Campoplex cf. dubitator Horst., Liotryphon sp., Lissonotus sp., Pimpla spuria (Grav.), Pimpla turionellae (L.), and some others, and a braconid (Apanteles britannicus Wilk.) in 2001–2002.     

First record of metacestodes of Mesocestoides sp. in the common starling (Sturnus vulgaris) in Europe, with an 18S rDNA characterisation of the isolate

Metacestodes of Mesocestoides sp. were recorded from Sturnus vulgaris (Passeriformes: Stumidae) in the Czech Republic in April 2002. They were found in a cutaneous cyst and in the thoracic region of the body cavity of the bird. This is the first record of metacestodes of Mesocestoides sp. in this host species in Europe as well as the first finding of the formation of a cutaneous cyst provoked by this parasite. Additional specimens from Apodemus agrarius (Mammalia: Rodentia) from Bulgaria and Lacerta agilis (Reptilia: Squamata) from the Czech Republic were compared with that from S. vulgaris. Sequence data from the V4 variable region (18S rDNA) were used to compare genetic variability among these and previously characterized isolates of Mesocestoides spp. A number of distinct clades were recognized, with metacestodes from L. agilis showing the highest degree of relative divergence.     

Pathotypes of <Emphasis Type="Italic">Plasmodiophora brassicae</Emphasis> causing damage to oilseed rape in the Czech Republic and Poland

Winter oilseed rape (Brassica napus) is an important crop in the Czech Republic and Poland. Clubroot disease caused by the pathogen Plasmodiophora brassicae is a serious and still-growing problem for oilseed rape growers in both countries. The aim of this study was to evaluate the pathotype composition of P. brassicae populations from the Czech Republic and Poland, according to the three evaluation systems, and to determine soil inoculum loads for representative fields via traditional end-point PCR as well as quantitative PCR analysis. There were considerable differences between the populations of P. brassicae from both countries, and the number of pathotypes varied depending on the evaluation system and the threshold used to distinguish susceptible vs. resistant plant reactions. This is the first study comparing the effect of different thresholds. Using an index of disease (ID) of 25 % to distinguish susceptible vs. resistant reactions, there was a total of seven pathotypes identified based on the differentials of Williams, five with the system of Somé et al., and 18 with the European Clubroot Differential (ECD) set. However, based on a threshold of 50 %, there were nine pathotypes according to the evaluation system by Williams, four based on the differentials of Somé et al., and 15 with the ECD set. Changing of the thresholds led to the reclassification of some pathotypes. Several pathotypes were common in both countries. High amounts of pathogen DNA were found in many of the field soils analysed by quantitative PCR. There was a weak correlation between soil pH and infestation of P. brassicae for the Polish soils.     

Characterization of Longidorus helveticus (Nematoda: Longidoridae) from the Czech Republic

Longidorus helveticus was found at two out of 285 sampling sites for the first time in the Czech Republic. Females, males and juvenile stages were analyzed morphologically and morphometrically. The morphological identification of samples was verified by polymerase chain reaction using a species specific primer. Four markers of ribosomal DNA (18S, ITS1, ITS2, D2-D3 expansion segments of 28S rRNA) and two markers of mitochondrial DNA (cox1 and nad4) were sequenced and analyzed and compared with published gene sequences of other populations of L. helveticus. The partial mitochondrial cytochrome c-oxidase subunit 1 gene and partial nicotinamide dehydrogenase subunits 4 gene showed relatively high genetic variation within the species compared with ribosomal DNA markers.