Coinfection of Babesia and Borrelia in the Tick Ixodes ricinus-A Neglected Public Health Issue in Europe?

Ixodes ricinus nymphs and adults removed from humans, and larvae and nymphs from birds, have been analysed for infection with Babesia species and Borrelia species previously in separately published studies. Here, we use the same data set to explore the coinfection pattern of Babesia and Borrelia species in the ticks. We also provide an overview of the ecology and potential public health importance in Sweden of I. ricinus infected both with zoonotic Babesia and Borrelia species. Among 1952 nymphs and adult ticks removed from humans, 3.1% were PCR-positive for Babesia spp. Of these Babesia-positive ticks, 43% were simultaneously Borrelia-positive. Among 1046 immatures of I. ricinus removed from birds, 2.5% were Babesia-positive, of which 38% were coinfected with Borrelia species. This study shows that in I. ricinus infesting humans or birds in Sweden, potentially zoonotic Babesia protozoa sometimes co-occur with human-pathogenic Borrelia spp. Diagnostic tests for Babesia spp. infection are rarely performed in Europe, and the medical significance of this pathogen in Europe could be underestimated.

Temporal chromatin accessibility changes define transcriptional states essential for osteosarcoma metastasis.

The metastasis-invasion cascade describes the series of steps required for a cancer cell to successfully spread from its primary tumor and ultimately grow within a secondary organ. Despite metastasis being a dynamic, multistep process, most omics studies to date have focused on comparing primary tumors to the metastatic deposits that define end-stage disease. This static approach means we lack information about the genomic and epigenomic changes that occur during the majority of tumor progression. One particularly understudied phase of tumor progression is metastatic colonization, during which cells must adapt to the new microenvironment of the secondary organ. Through temporal profiling of chromatin accessibility and gene expression in vivo, we identify dynamic changes in the epigenome that occur as osteosarcoma tumors form and grow within the lung microenvironment. Furthermore, we show through paired in vivo and in vitro CRISPR drop-out screens and pharmacological validation that the upstream transcription factors represent a class of metastasis-specific dependency genes. While current models depict lung colonization as a discrete step within the metastatic cascade, our study shows it is a defined trajectory through multiple epigenetic states, revealing new therapeutic opportunities undetectable with standard approaches.

Characteristics of Human Babesiosis in Europe.

One of the Editor's choice articles in 2021 published in Pathogens was a review of human babesiosis in Europe [...].

The biology of Ixodes ricinus with emphasis on its ecology.

Prior to its identification as the vector of Lyme borreliosis spirochaetes in Europe in 1983, interest in Ixodes ricinus (L.) was moderate and mainly concerned the transmission of pathogens to farm animals and of tick-borne encephalitis virus to humans. The situation now is very different, and more papers have been published on I. ricinus than on any other ixodid tick species. However, this large literature is scattered and in recent years has become dominated by the molecular detection and characterization of the many pathogens that I. ricinus transmits. Several decades have now elapsed since a review addressing its basic biology and ecology appeared, and the present publication seeks to present basic aspects of its biology and ecology that are related to its role as a vector of disease agents, including its life cycle, feeding behaviour, host relations, survival off the host, and the impact of weather and climate.

Babesia and Human Babesiosis.

Babesia is a genus of intraerythrocytic protozoan parasites belonging to the exclusively parasitic phylum Apicomplexa [...].

The Prevalence and Genetic Diversity of Babesia divergens in Ixodes ricinus Nymphs Collected from Farm- and Woodland Sites in Ireland.

The parasite, Babesia divergens causes redwater fever in cattle and a rare, albeit life-threatening disease in humans. In Ireland, B. divergens has always been considered an important pathogen as the high incidence of redwater fever precluded areas of the country from cattle farming. Moreover a relatively large proportion of human cases were reported here. Red deer (Cervus elaphus), which often harbour babesias that are genetically very similar (if not identical) to B. divergens, are quite widespread. In this study 1369 nymphal Ixodes ricinus ticks collected from various habitats were screened for the presence of B. divergens using TaqMan followed by conventional nested PCR. Fragments of the 18S rRNA gene locus (560 bp) were compared against published Irish B. divergens isolates from cattle, humans and red deer. Overall just 1% of I. ricinus nymphs were infected with B. divergens, with similar infection rates in ticks collected from farm- and woodland. Most (90%) 18S rRNA gene fragments derived from woodland ticks were 100% identical to published sequences from cattle and humans. One differed by a single nucleotide polymorphism (SNP) as did two isolates from ticks collected in bogland. Two isolates derived from nymphs collected in farmland differed by 2 and 4 SNPs respectively.

Babesia and Theileria Identification in Adult Ixodid Ticks from Tapada Nature Reserve, Portugal.

This study, conducted in a nature reserve in southern Portugal, investigated the frequency and diversity of tick-borne piroplasms in six species of adult ixodid ticks removed from 71 fallow deer (Dama dama) and 12 red deer (Cervus elaphus), collected over the period 2012-2019. The majority of 520 ticks were Ixodes ricinus (78.5%), followed by Rhipicephalus sanguineus sensu lato, Hyalomma lusitanicum, Haemaphysalis punctata, Dermacentor marginatus, and Ixodes hexagonus. The R. sanguineus ticks collected from the deer were clearly exophilic, in contrast to the endophilic species usually associated with dogs. Four tick-borne piroplasms, including Theileria spp., and the zoonotic species, Babesia divergens and Babesia microti, were detected. B. divergens 18S rDNA, identical to that of the bovine reference strain U16370 and to certain strains from red deer, was detected in I. ricinus ticks removed from fallow deer. The sporadic detection of infections in ticks removed from the same individual hosts suggests that the piroplasms were present in the ticks rather than the hosts. Theileria sp. OT3 was found in I. ricinus and, along with T. capreoli, was also detected in some of the other tick species. The natural vector and pathogenic significance of this piroplasm are unknown.

Ticks, Human Babesiosis and Climate Change.

The effects of current and future global warming on the distribution and activity of the primary ixodid vectors of human babesiosis (caused by Babesia divergens, B. venatorum and B. microti) are discussed. There is clear evidence that the distributions of both Ixodes ricinus, the vector in Europe, and I. scapularis in North America have been impacted by the changing climate, with increasing temperatures resulting in the northwards expansion of tick populations and the occurrence of I. ricinus at higher altitudes. Ixodes persulcatus, which replaces I. ricinus in Eurasia and temperate Asia, is presumed to be the babesiosis vector in China and Japan, but this tick species has not yet been confirmed as the vector of either human or animal babesiosis. There is no definite evidence, as yet, of global warming having an effect on the occurrence of human babesiosis, but models suggest that it is only a matter of time before cases occur further north than they do at present.

Human Babesiosis in Europe.

Babesiosis is attracting increasing attention as a worldwide emerging zoonosis. The first case of human babesiosis in Europe was described in the late 1950s and since then more than 60 cases have been reported in Europe. While the disease is relatively rare in Europe, it is significant because the majority of cases present as life-threatening fulminant infections, mainly in immunocompromised patients. Although appearing clinically similar to human babesiosis elsewhere, particularly in the USA, most European forms of the disease are distinct entities, especially concerning epidemiology, human susceptibility to infection and clinical management. This paper describes the history of the disease and reviews all published cases that have occurred in Europe with regard to the identity and genetic characteristics of the etiological agents, pathogenesis, aspects of epidemiology including the eco-epidemiology of the vectors, the clinical courses of infection, diagnostic tools and clinical management and treatment.

What do we still need to know about Ixodes ricinus?

In spite of many decades of intensive research on Ixodes ricinus, the castor bean tick of Europe, several important aspects of its basic biology remain elusive, such as the factors determining seasonal development, tick abundance and host specificity, and the importance of water management. Additionally, there are more recent questions about the geographical diversity of tick genotypes and phenotypes, the role of migratory birds in the ecoepidemiology of I. ricinus, the importance of protective immune responses against I. ricinus, particularly in the context of vaccination, and the role of the microbiome in pathogen transmission. Without more detailed knowledge of these issues, it is difficult to assess the likely effects of changes in climate and biodiversity on tick distribution and activity, to predict potential risks arising from new and established tick populations and I. ricinus-borne pathogens, and to improve prevention and control measures. This review aims to discuss the most important outstanding questions against the backdrop of the current state of knowledge of this important tick species.

Four-Dimensional Characterization of the Babesia divergens Asexual Life Cycle, from the Trophozoite to the Multiparasite Stage.

Babesia is an apicomplexan parasite of significance that causes the disease known as babesiosis in domestic and wild animals and in humans worldwide. Babesia infects vertebrate hosts and reproduces asexually by a form of binary fission within erythrocytes/red blood cells (RBCs), yielding a complex pleomorphic population of intraerythrocytic parasites. Seven of them, clearly visible in human RBCs infected with Babesia divergens, are considered the main forms and named single, double, and quadruple trophozoites, paired and double paired pyriforms, tetrad or Maltese Cross, and multiparasite stage. However, these main intraerythrocytic forms coexist with RBCs infected with transient parasite combinations of unclear origin and development. In fact, little is understood about how Babesia builds this complex population during its asexual life cycle. By combining cryo-soft X-ray tomography and video microscopy, main and transitory parasites were characterized in a native whole cellular context and at nanometric resolution. The architecture and kinetics of the parasite population was observed in detail and provide additional data to the previous B. divergens asexual life cycle model that was built on light microscopy. Importantly, the process of multiplication by binary fission, involving budding, was visualized in live parasites for the first time, revealing that fundamental changes in cell shape and continuous rounds of multiplication occur as the parasites go through their asexual multiplication cycle. A four-dimensional asexual life cycle model was built highlighting the origin of several transient morphological forms that, surprisingly, intersperse in a chronological order between one main stage and the next in the cycle.IMPORTANCE Babesiosis is a disease caused by intraerythrocytic Babesia parasites, which possess many clinical features that are similar to those of malaria. This worldwide disease is increasing in frequency and geographical range and has a significant impact on human and animal health. Babesia divergens is one of the species responsible for human and cattle babesiosis causing death unless treated promptly. When B. divergens infects its vertebrate hosts, it reproduces asexually within red blood cells. During its asexual life cycle, B. divergens builds a population of numerous intraerythrocytic (IE) parasites of difficult interpretation. This complex population is largely unexplored, and we have therefore combined three- and four-dimensional imaging techniques to elucidate the origin, architecture, and kinetics of IE parasites. Unveiling the nature of these parasites has provided a vision of the B. divergens asexual cycle in unprecedented detail and is a key step to develop control strategies against babesiosis.

Update on the presence of Ixodes ricinus at the western limit of its range and the prevalence of Borrelia burgdorferi sensu lato.

It is often suggested that due to climate and environmental policy changes, the risk from tick-borne disease is increasing, particularly at the geographical limits of the vector distribution. Our project aimed to determine whether this was true for the risk of Lyme borreliosis in Ireland which is the western-most limit of Ixodes ricinus, the European vector of Borrelia burgdorferi sensu lato. The availability of a historical data set of tick infection rates compiled in the 1990s represented a unique opportunity as it provided a baseline against which current data could be compared. Following construction of a spatial predictive model for the presence and absence of I. ricinus based on data from 491 GPS locations visited between 2016 and 2019, 1404 questing nymphs from 27 sites were screened for the presence of Borrelia spp. using a TaqMan PCR aimed at the 23S rRNA gene sequence. All positive ticks were further analysed by nested PCR amplification and sequence analysis of the 5 S-23 S intergenic spacer. The model indicated that areas with the highest probability of tick presence were mostly located along the western seaboard and the Shannon and Erne river catchments, coinciding with historical high incidence areas of bovine babesiosis, while the infection rate of questing nymphs with B. burgdorferi s.l. and the prevalence of the various genospecies have remained surprisingly stable over the last 3 decades. Clear communication of the potential disease risk arising from a tick bite is essential in order to allay undue concerns over tick-borne diseases among the general public.

PsychoPy2: Experiments in behavior made easy.

PsychoPy is an application for the creation of experiments in behavioral science (psychology, neuroscience, linguistics, etc.) with precise spatial control and timing of stimuli. It now provides a choice of interface; users can write scripts in Python if they choose, while those who prefer to construct experiments graphically can use the new Builder interface. Here we describe the features that have been added over the last 10 years of its development. The most notable addition has been that Builder interface, allowing users to create studies with minimal or no programming, while also allowing the insertion of Python code for maximal flexibility. We also present some of the other new features, including further stimulus options, asynchronous time-stamped hardware polling, and better support for open science and reproducibility. Tens of thousands of users now launch PsychoPy every month, and more than 90 people have contributed to the code. We discuss the current state of the project, as well as plans for the future.

Vectors of Babesiosis.

Babesiosis, caused by piroplasmid protozoans in the genus Babesia, is arguably the most important vector-borne disease of livestock and companion animals and is growing in importance as a zoonosis. Ixodid ticks were identified as vectors more than a hundred years ago, but the particular tick species transmitting some significant pathogens are still unknown. Moreover, it is only recently that the complexity of the pathogen-tick relationship has been revealed as a result of studies enabled by gene expression and RNA interference methodology. In this article, we provide details of demonstrated and incriminated vectors, maps of the current knowledge of vector distribution, a summary of established features of the pathogen life cycle in the vector, and an outline of molecular research on pathogen-tick relationships. The article concludes with a discussion of vector ecology and disease epidemiology in a global-change context and with suggestions for future research.

Kinetics of the invasion and egress processes of Babesia divergens, observed by time-lapse video microscopy.

Based on confocal fluorescence and bright field video microscopy, we present detailed observations on the processes of invasion and egress of erythrocytes by the apicomplexan parasite Babesia divergens. Time-lapse images reveal numerous unexpected findings associated with the dynamics of B. divergens and its ability to manipulate the erythrocyte during both processes in its asexual cycle under in vitro conditions. Despite the speed at which these processes occur and the small size of the parasite, we capture infective merozoites moving vigorously and causing striking deformations in the erythrocyte's plasma membrane during an active invasion. We also observed intraerythrocytic dynamic stages as paired pyriforms, double paired pyriforms, tetrads, unattached pyriform sister cells and multiple parasite stages resulting in the release of large numbers of merozoites over a short period. Of considerable interest is that time-lapse images reveal a novel mechanism of egress used by B. divergens to exit the human erythrocyte. The release occurs when B. divergens parasites establish contacts with the plasma membrane of the erythrocyte from within, before exiting the cell. Visualization and analysis of the images enabled us to obtain useful information and broaden our knowledge of complex and crucial events involved with parasitisation of human erythrocytes by B. divergens.

Ecosystem change and zoonoses in the Anthropocene.

Changes in land use, animal populations and climate, primarily due to increasing human populations, drive the emergence of zoonoses. Force of infection (FOI), which for these diseases is a measure of the ease with which a pathogen reaches the human population, can change with specific zoonoses and context. Here, we outline three ecosystem categories-domestic, peridomestic and sylvatic, where disease ecology alters the FOI of specific zoonoses. Human intervention is an overriding effect in the emergence of zoonoses; therefore, we need to understand the disease ecology and other influencing factors of pathogens and parasites that are likely to interact differently within ecological and cultural contexts. Planning for One Health and community ecology, such as an ecological impact assessment, is required to prepare and manage the emergence and impact of zoonoses in the Anthropocene.

Misdiagnosis of Babesiosis as Malaria, Equatorial Guinea, 2014.

We report a case of babesiosis, caused by Babesia microti, in a missionary who worked in Equatorial Guinea but also visited rural Spain. The initial diagnosis, based on clinical features and microscopy, was malaria. The patient's recovery was delayed until she received appropriate treatment for babesiosis.

Research on Piroplasmorida and other tick-borne agents: Are we going the right way?

The use of molecular diagnostic techniques in the study of ticks and tick-borne agents has increased dramatically in the last few decades, and while conferring the advantages of rapidity, sensitivity and accuracy, they can also be misused by the research community, who have shown an increasing overreliance on the data they produce, often completely abandoning more traditional approaches such as microscopy and serology. In this short review examples are provided of 'findings' that are apparently at odds with well-established eco-epidemiological knowledge, ranging from descriptions of new species of pathogens to their detection in new hosts and ticks, often immediately incriminated as new reservoir hosts and vector ticks, respectively. Most examples have been taken from the Piroplasmorida or "piroplasm" species as they are often called. Researchers are urged to utilize molecular methods with as much awareness as possible of the system or systems they are investigating, and journal editors and their teams of reviewers should be encouraged to adopt a more critical approach to papers that contain data based solely on molecular methods without due regard to the underlying biology. Explanations for such findings should be offered and discussed.