A mézelő méh (Apis mellifera L.) vírusfertőzéseinek és a méhpatogén vírusok molekuláris szerkezetének tanulmányozása = Investigations on the viral infections of the honey bee (Apis mellifera L.) and on the molecular structure of bee pathogens

A mézelő méh vírusainak vizsgálatára irányuló kutatások keretében polimeráz láncreakcióra (PCR) alapozott felmérő vizsgálatokat végeztünk magyarországi és ausztriai méhészetekben a hat legfontosabb vírus előfordulási gyakoriságának megismerésére. Elsőként mutattuk ki a méhek szárnydeformitását okozó vírus, a fekete anyabölcső vírus, a lárvatömlősödés vírus és a krónikus méhbénulás vírus előfordulását hazai és ausztriai méhészetekben. Eltéréseket tapasztaltunk a vírusok hazai és külföldi előfordulási gyakorisága között. Filogenetikai vizsgálatokat végeztünk a fekete anyabölcső vírus közép-európai törzseinek bevonásával és feltártuk a vírus különböző genomterületeinek változékonyságát. A méhek szárnydeformitását okozó vírus világszerte gyűjtött törzseinek filogenetikai összehasonlítása során nagyon közeli rokonságot tapasztaltunk a vizsgált szekvenciák között, amely a vírus közelmúltbeli gyors terjedésére utal. Ez a terjedés valószínűleg összefüggésben áll a Varroa destructor atka elterjedésével, mivel ez a parazita hatékony vektora a vírusnak. Meghatároztuk a heveny méhbénulás vírus egy magyarországi és egy lengyelországi törzsének teljes genomszekvenciáját. A zab levéltetű (Rhopalosiphum padi) vírusát mutattuk ki méh eredetű mintákból. Folytattuk a hazai méhekből korábban kimutatott, a heveny méhbénuláshoz és a Kashmir méhvírushoz hasonló vírus genetikai jellemzését. Három méhvírus egyidejű kimutatására alkalmas multiplex RT-PCR alapú diagnosztikai módszert fejlesztettünk ki. | The project was focused on the investigations of honeybee viruses. Polymerase chain reaction (PCR)-based surveillance was performed on bee samples from Hungarian and Austrian apiaries to record the incidence of the six most important bee viruses. We have detected the deformed wing virus (DWV), the black queen cell virus (BQCV), the sacbrood virus, and the chronic bee paralysis virus for the first time in these countries. We found considerable differences in the prevalence of these viruses in different countries. Phylogenetic analyses were made on central European BQCV strains, and the variability of the different genome regions was recorded. By the genetic analysis of DWV strains collected worldwide, we detected close genetic relationship between the strains, which indicates a recent spread of the virus. This is probably in connection with the global spread of Varroa destructor, because this parasitic mite is an efficient vector of the virus. We have determined the complete genome sequence of a Hungarian and a Polish acute bee paralysis virus (ABPV) strain. We have detected the bird cherry aphid (Rhopalosiphum padi) virus in bee samples. We carried on the genetic characterization of a bee virus, which was detected in Hungary, and which is closely related to the ABPV and the Kashmir bee virus. We have developed a multiplex RT-PCR assay for the simultaneous detection of three viruses in honeybee samples

Studies of ticks (Acari: Ixodidae) and tick-borne pathogens of dogs in Hungary

In Europe, the number of reports on canine tick-borne diseases has increased in the past few years. In Hungary, we have had very limited information concerning tick infestation and tick-borne pathogens of dogs. For these reasons, we started to study the tick species and tick-borne pathogens infecting dogs in our country. Based on morphological studies, a figured practical identification key has been designed for the sixteen hard tick species which have been found on dogs in Europe. The simplicity of this key can help veterinarians and zoologists in tick identification. In 29 veterinary clinics from six districts of Budapest and 13 counties, 1779 tick specimens were collected from 606 dogs. Most hosts were usually infested with a single female and very few of them had many ticks. The most preferred sites of tick attachment in decreasing order were head, neck and legs. Ixodes ricinus and Dermacentor reticulatus were the most common species. Ixodes canisuga, Haemaphysalis concinna, Ixodes hexagonus, Ixodes acuminatus and Dermacentor marginatus were also found. New data have been provided about the geographical distribution of Dermacentor reticulatus, because the specimens of this species were collected in north-eastern and south-eastern parts of the country too where they had not been found before. Field collections in 31 locations provided new data on the geographical and seasonal occurrence of I. ricinus, D. reticulatus and other tick species as well. The occurrence of small canine piroplasms in two dogs was described for the first time in Hungary. These were autochtonous infestations but we need further investigations to know the species, occurrence, vector and origin of this pathogen. The subspecies Babesia canis canis was identified to be the causative agent of babesiosis caused by large Babesia sp. in dogs using molecular biological methods. It was also proven with molecular methods that the geographical distribution of canine babesiosis is larger in the country than it has been previously known. Babesia DNA was detected in free-living and engorged D. reticulatus females for the first time in the country. Presence of B. canis canis in engorged D. reticulatus specimens removed from dogs was also demonstrated with molecular methods. Molecular evidence was found for the presence of Borrelia sp. in free-living and engorged I. ricinus females for the first time in Hungary. Three species, B. burgdorferi s.s., B. afzelii and B. garinii were identified with sequence analysis which are pathogenic to both dogs and humans

Analysis of a novel RNA virus in a wild northern white-breasted hedgehog (Erinaceus roumanicus)

Tombusviruses are generally considered plant viruses. A novel tombus-/carmotetravirus-like RNA virus was identified in a faecal sample and blood and muscle tissues from a wild northern white-breasted hedgehog (Erinaceus roumanicus). The complete genome of the virus, called H14-hedgehog/2015/HUN (GenBank accession number MN044446), is 4,118 nucleotides in length with a readthrough stop codon of type/group 1 in ORF1 and lacks a poly(A) tract at the 3' end. The predicted ORF1-RT (RdRp) and the capsid proteins had low (31-33%) amino acid sequence identity to unclassified tombus-/noda-like viruses (Hubei tombus-like virus 12 and Beihai noda-like virus 10), respectively, discovered recently in invertebrate animals. An in vivo experimental plant inoculation study showed that an in vitro-transcribed H14-hedgehog/2015/HUN viral RNA did not replicate in Nicotiana benthamiana, Chenopodium quinoa, or Chenopodium murale, the most susceptible hosts for plant-origin tombusviruses

Eco-epidemiology of Borrelia miyamotoi and Lyme borreliosis spirochetes in a popular hunting and recreational forest area in Hungary

BACKGROUND: Borrelia miyamotoi, the newly discovered human pathogenic relapsing fever spirochete, and Borrelia burgdorferi sensu lato are maintained in natural rodent populations. The aim of this study was to investigate the natural cycle of B. miyamotoi and B. burgdorferi s.l. in a forest habitat with intensive hunting, forestry work and recreational activity in Southern Hungary. METHODS: We collected rodents with modified Sherman-traps during 2010–2013 and questing ticks with flagging in 2012. Small mammals were euthanized, tissue samples were collected and all ectoparasites were removed and stored. Samples were screened for pathogens with multiplex quantitative real-time polymerase chain reaction (qPCR) targeting a part of flagellin gene, then analysed with conventional PCRs and sequencing. RESULTS: 177 spleen and 348 skin samples of six rodent species were individually analysed. Prevalence in rodent tissue samples was 0.2 % (skin) and 0.5 % (spleen) for B. miyamotoi and 6.6 % (skin) and 2.2 % (spleen) for B. burgdorferi s.l. Relapsing fever spirochetes were detected in Apodemus flavicollis males, B. burgdorferi s.l. in Apodemus spp. and Myodes glareolus samples. Borrelia miyamotoi was detected in one questing Ixodes ricinus nymph and B. burgdorferi s.l in nymphs and adults. In the ticks removed from rodents DNA amplification of both pathogens was successful from I. ricinus larvae (B. miyamotoi 5.6 %, B. burgdorferi s.l. 11.1 %) and one out of five nymphs while from Ixodes acuminatus larvae, and nymph only B. burgdorferi s.l. DNA was amplified. Sequencing revealed B. lusitaniae in a questing I. ricinus nymph and altogether 17 B. afzelii were identified in other samples. Two Dermacentor marginatus engorged larva pools originating from uninfected hosts were also infected with B. afzelii. CONCLUSIONS: This is the first report of B. miyamotoi occurrence in a natural population of A. flavicollis as well as in Hungary. We provide new data about circulation of B. burgdorferi s.l. in rodent and tick communities including the role of I. acuminatus ticks in the endophilic pathogen cycle. Our results highlight the possible risk of infection with relapsing fever and Lyme borreliosis spirochetes in forest habitats especially in the high-risk groups of hunters, forestry workers and hikers

The 3P Framework: A Comprehensive Approach to Coping with the Emerging Infectious Disease Crisis

The COVID-19 pandemic is the latest example of the profound socioeconomic impact of the emerging infectious disease (EID) crisis. Current health security measures are based on a failed evolutionary paradigm that presumes EID is rare and cannot be predicted because emergence requires the prior evolution of novel genetic capacities for colonizing a new host. Consequently, crisis response through preparation for previously emerged diseases and palliation following outbreaks have been the only health security options, which have become unsustainably expensive and unsuccessful. The Stockholm paradigm (SP) is an alternative evolutionary framework that suggests host changes are the result of changing conditions that bring pathogens into contact with susceptible hosts, with novel genetic variants arising in the new host after infection. Host changes leading to EID can be predicted because preexisting capacities for colonizing new hosts are highly specific and phylogenetically conservative. This makes EID prevention through limiting exposure to susceptible hosts possible. The DAMA (Document, Assess, Monitor, Act) protocol is a policy extension of the SP that can both prevent and mitigate EID by enhancing traditional efforts through adding early warning signs and predicting transmission dynamics. Prevention, preparation, and palliation compose the 3P framework, a comprehensive plan for reducing the socioeconomic impact of EID. This article has been produced in support of and with appreciation for the efforts by Gábor Földvári of the Institute of Evolution, Centre for Ecological Research, and the Centre for Eco-Epidemiology, National Laboratory for Health Security (both located at 1121 Budapest, Konkoly-Thege Miklós út 29-33, Hungary). Through his untiring efforts, team building, and leadership, he has secured the first EU-wide team research grant. This work was supported by the National Research, Development and Innovation Office in Hungary (RRF-2.3.1-21-2022-00006) and the COST Action CA21170 “Prevention, anticipation and mitigation of tick-borne disease risk applying the DAMA protocol (PRAGMATICK),” which represent the first funded efforts to apply the principles of the DAMA protocol

Vertical transmission of Bartonella schoenbuchensis in Lipoptena cervi.

BACKGROUND Lipoptena cervi (Diptera: Hippoboscidae) is a hematophagous ectoparasite of cervids, which is considered to transmit pathogens between animals and occasionally to humans. The principal life stage that is able to parasitize new hosts is a winged ked that just emerged from a pupa. To facilitate efficient transmission of pathogens between hosts, vertical transmission from female deer keds to their offspring is necessary. We investigated vertical transmission of several vector-borne pathogens associated with cervids. METHODS Deer keds from several locations in Hungary were collected between 2009 and 2012. All life stages were represented: winged free-ranging adults, wingless adults collected from Capreolus capreolus and Cervus elaphus, developing larvae dissected from gravid females, and fully developed pupae. The presence of zoonotic pathogens was determined using qPCR or conventional PCR assays performed on DNA lysates. From the PCR-positive lysates, a gene fragment was amplified and sequenced for confirmation of pathogen presence, and/or pathogen species identification. RESULTS DNA of Bartonella schoenbuchensis was found in wingless males (2%) and females (2%) obtained from Cervus elaphus, dissected developing larvae (71%), and free-ranging winged males (2%) and females (11%). DNA of Anaplasma phagocytophilum and Rickettsia species was present in L. cervi adults, but not in immature stages. DNA of Candidatus Neoehrlichia mikurensis was absent in any of the life stages of L. cervi. CONCLUSIONS B. schoenbuchensis is transmitted from wingless adult females to developing larvae, making it very likely that L. cervi is a vector for B. schoenbuchensis. Lipoptena cervi is probably not a vector for A. phagocytophilum, Rickettsia species, and Candidatus N. mikurensis