High-resolution epidemiological landscape from ~290,000 SARS-CoV-2 genomes from Denmark.

Vast amounts of pathogen genomic, demographic and spatial data are transforming our understanding of SARS-CoV-2 emergence and spread. We examined the drivers of molecular evolution and spread of 291,791 SARS-CoV-2 genomes from Denmark in 2021. With a sequencing rate consistently exceeding 60%, and up to 80% of PCR-positive samples between March and November, the viral genome set is broadly whole-epidemic representative. We identify a consistent rise in viral diversity over time, with notable spikes upon the importation of novel variants (e.g., Delta and Omicron). By linking genomic data with rich individual-level demographic data from national registers, we find that individuals aged  < 15 and  > 75 years had a lower contribution to molecular change (i.e., branch lengths) compared to other age groups, but similar molecular evolutionary rates, suggesting a lower likelihood of introducing novel variants. Similarly, we find greater molecular change among vaccinated individuals, suggestive of immune evasion. We also observe evidence of transmission in rural areas to follow predictable diffusion processes. Conversely, urban areas are expectedly more complex due to their high mobility, emphasising the role of population structure in driving virus spread. Our analyses highlight the added value of integrating genomic data with detailed demographic and spatial information, particularly in the absence of structured infection surveys.

Communicating and disseminating One Health: successes of the One Health European Joint Programme.

The application of a One Health approach recognizes that human health, animal health, plant health and ecosystem health are intrinsically connected. Tackling complex challenges associated with foodborne zoonoses, antimicrobial resistance, and emerging threats is imperative. Therefore, the One Health European Joint Programme was established within the European Union research programme Horizon 2020. The One Health European Joint Programme activities were based on the development and harmonization of a One Health science-based framework in the European Union (EU) and involved public health, animal health and food safety institutes from almost all EU Member States, the UK and Norway, thus strengthening the cooperation between public, medical and veterinary organizations in Europe. Activities including 24 joint research projects, 6 joint integrative projects and 17 PhD projects, and a multicountry simulation exercise facilitated harmonization of laboratory methods and surveillance, and improved tools for risk assessment. The provision of sustainable solutions is integral to a One Health approach. To ensure the legacy of the work of the One Health European Joint Programme, focus was on strategic communication and dissemination of the outputs and engagement of stakeholders at the national, European and international levels.

Epidemic of parvovirus B19 and disease severity in pregnant people, Denmark, January to March 2024.

We report an epidemic of parvovirus B19 infections in Denmark during the first quarter of 2024, with a peak incidence 3.5 times higher than during the most recent epidemic in 2017. In total, 20.1% (130/648) of laboratory-confirmed cases were pregnant. Severe adverse outcomes were observed among 12.3% (16/130) of pregnant people and included foetal anaemia, foetal hydrops and miscarriage. Parvovirus B19 infection is not systematically monitored, but a national laboratory-based surveillance system is currently being established in Denmark.

Global prevalence of Toxoplasma gondii in birds: A systematic review and meta-analysis.

Among the potential animal reservoirs of the zoonotic parasite T. gondii, birds have received relatively little attention. This systematic review and meta-analysis aimed to assess the global status and to provide an overview of the epidemiology of T. gondii infection in birds. The standard protocol of preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines were followed. Scopus, PubMed, Web of Science, Science Direct, ProQuest, and Google Scholar were searched for relevant publications from January 1990, to March 2024. All peer-reviewed original research articles describing the prevalence of T. gondii in birds were included. Inclusion and exclusion criteria were applied, and both direct and indirect detection were considered. The point estimates and 95% confidence intervals were calculated using the meta-package in R (version 3.6.1). The variance between studies (heterogeneity) was quantified by the I2 index. Finally, 258 articles (including 380 datasets) were eligible for inclusion in the systematic review and meta-analysis. The global pooled prevalence was 24% (21 - 26%). The highest prevalence of T. gondii was observed in buzzards (52%, 34 - 70%), turkeys (31%, 17 - 46%), and chickens (30%, 26 - 34%). The present study provides a comprehensive view of the global prevalence of T. gondii in birds.

Heartworm disease in domestic dogs in Estonia: indication of local circulation of the zoonotic parasite Dirofilaria immitis farther north than previously reported.

BACKGROUND: The mosquito-borne zoonotic parasite Dirofilaria immitis continues to spread northwards in Europe. This parasite can cause potentially life-threatening heartworm disease in dogs and pulmonary dirofilariasis in humans and is, therefore, a major health concern in both the veterinary medicine and human medical fields. This is the first report of D. immitis infections and heartworm disease in the Baltic country Estonia. METHODS: Data on canine D. immitis infections and heartworm disease were collected from the electronic patient records database of the Small Animal Clinic of Estonian University of Life Sciences, the only university clinic in Estonia. The patient records of dogs with confirmed diagnosis of D. immitis infection or heartworm disease were reviewed and summarised. RESULTS: Six dogs had been diagnosed with confirmed D. immitis infection or heartworm disease at the university clinic in 2021-2022. The confirmed diagnoses had been reached following international guidelines, based on a combination of different tests. Molecular confirmation of the parasite species had not been performed. Two of the dogs had been imported while four had no travel history outside of the country. CONCLUSIONS: Four of the dogs with a confirmed D. immitis infection or heartworm disease had no history of being imported or travelling outside of the country, indicating autochthonous infections and, consequently, local circulation of the parasite in Estonia. These findings represent the new northernmost autochthonous cases of D. immitis infection and canine heartworm disease reported in the European Union.

Veterinarians and zoonotic pathogens, infections and diseases - questionnaire study and case series, Finland.

BACKGROUND: Veterinarians are at risk for numerous zoonotic infections. In this paper, we summarise descriptions of zoonotic infections from a questionnaire study and a series of work-related zoonotic cases, aiming to add to the knowledge on occupational zoonotic risks of veterinarians. METHODS: We collected data on zoonotic infections contracted by veterinarians in Finland in two studies:1) using a questionnaire in 2009, and 2) inviting veterinarians who had encountered an occupational zoonosis to report it in structured interviews in 2019. RESULTS AND CONCLUSIONS: In the questionnaire study in 2009, of 306 veterinarians several reported zoonotic bacterial skin infections (12%), dermatophytosis (ringworm; 4.2%), virus infections (3.9%), bacterial gastroenteritis (3.3%), other bacterial zoonoses (2.3%), and parasitic infections/infestations (2.3%). In the 2019 interviews, 16 occupational zoonosis cases were reported. Of them, seven were selected to the case series. The selected cases included Capnocytophaga canimorsus sepsis following a dog bite, cryptosporidiosis after a contact with calves, cutaneous listeriosis following calving assistance, Salmonella gastroenteritis contracted at laboratory, Trichophyton dermatophytosis after equine contact, Bacillus anthracis exposure at necropsy, and exposure to rabies through a horse bite. In four of the seven cases, the veterinarian disagreed or strongly disagreed with having had good knowledge of the zoonosis before the incident. The results from the questionnaire study and the case series illustrate the variety of zoonotic pathogens that veterinarians may encounter. There is a need to improve the occupational health of veterinarians and to increase awareness in the occupational health sector. We encourage addressing this need using a One Health approach.

Detection of zoonotic Cryptosporidium spp. in small wild rodents using amplicon-based next-generation sequencing.

Rodents may serve as reservoirs of zoonotic species of Cryptosporidium; however, data from molecular surveys in support of this hypothesis are still scarce. In this study, we screened faeces and rectal content from murid and cricetid rodents (N = 58) caught around three farms in Zealand, Denmark, for Cryptosporidium spp. by amplicon-based next-generation sequencing (NGS) of ribosomal genes. Selected samples were further examined using nested conventional PCR targeting SSU rRNA, gp60, and actin genes. Cryptosporidium-specific DNA was identified in 40/58 (69%) samples, and in 12 (30%) of the 40 positive animals, mixed cryptosporidial infections were observed. Cryptosporidium ditrichi was the species most commonly identified, found in 28 (48%) of the animals. Cryptosporidium parvum was identified in 4 (7%) of the animals, all of which were co-infected with C. ditrichi. The present study is the first to utilize NGS-based screening for Cryptosporidium species in wild rodents. Moreover, it is the first study to provide molecular data on Cryptosporidium in rodents sampled in Denmark and to detect DNA of C. ditrichi in Mus musculus, Myodes glareolus, and Microtus agrestis. The NGS approach was successfully applied to yield new knowledge, and the results showed that zoonotic species of Cryptosporidium are common in murid and cricetid rodents in Zealand, Denmark.

Seroprevalence of Toxoplasma gondii in wild boars (Sus scrofa) hunted in Ukraine.

Toxoplasma gondii is an important zoonotic parasite worldwide, but it has received limited attention in Ukraine. A seroepidemiological study was conducted and samples from 452 wild boars that had been hunted in 2006-2011 in 23 of the 25 regions of Ukraine were tested to estimate T. gondii seroprevalence. A locally available commercial enzyme-linked immunosorbent assay (ELISA) was used for the investigation. Additionally, we tested 92 of the sera using a widely used commercial multi-species ELISA and an indirect immunofluorescence antibody test (IFAT). With the locally available ELISA, 35 of the 452 wild boars tested positive, yielding a seroprevalence estimate of 7.7% (95% confidence interval 5.5-10.5). The seropositive wild boars originated from eight of the regions. Using the majority criteria, 10/92 samples tested using both ELISAs and the IFAT were considered positive, yielding an estimated seroprevalence of 10.9% within the subset of samples. The highest seroprevalence was observed in wild boars hunted in Luhans'k (30.0%), Odesa (17.7%) and Kharkiv (12.7%). Seroprevalence was higher in older animals (13.3% for age group >12 months and 7.7% for age group ≤12 months). This is the first seroepidemiological study of T. gondii in wild boars in Ukraine. Assuming that seropositivity indicates presence of infectious parasites in the tissues, eating undercooked meat of wild boars hunted in Ukraine could be a potential source of infection to other hosts, including humans.

Mycoplasma pneumoniae epidemic in Denmark, October to December, 2023.

We report a surge of patients, especially children and adolescents, with respiratory disease caused by Mycoplasma pneumoniae in Denmark since October 2023. While the surge has reached an epidemic level, no impact on hospital capacity has been observed; only 14% (446/3,195) of cases, primarily adults, required hospitalisation. Macrolide resistance was detected in less than 2% of samples tested. Timely monitoring of hospitalisations linked to M. pneumoniae infections has been established to inform the healthcare system, decisionmakers and the public.

Mitochondrial genetic diversity and phylogenetic relationships of Echinococcus multilocularis in Europe.

The cestode Echinococcus multilocularis is the causative agent of alveolar echinococcosis, a fatal zoonotic parasitic disease of the northern hemisphere. Red foxes are the main reservoir hosts and, likely, the main drivers of the geographic spread of the disease in Europe. Knowledge of genetic relationships among E. multilocularis isolates at a European scale is key to understanding the dispersal characteristics of E. multilocularis. Hence, the present study aimed to describe the genetic diversity of E. multilocularis isolates obtained from different host species in 19 European countries. Based on the analysis of complete nucleotide sequences of the cob, atp6, nad2, nad1 and cox1 mitochondrial genes (4,968 bp), 43 haplotypes were inferred. Four haplotypes represented 62.56 % of the examined isolates (142/227), and one of these four haplotypes was found in each country investigated, except Svalbard, Norway. While the haplotypes from Svalbard were markedly different from all the others, mainland Europe appeared to be dominated by two main clusters, represented by most western, central and eastern European countries, and the Baltic countries and northeastern Poland, respectively. Moreover, one Asian-like haplotype was identified in Latvia and northeastern Poland. To better elucidate the presence of Asian genetic variants of E. multilocularis in Europe, and to obtain a more comprehensive Europe-wide coverage, further studies, including samples from endemic regions not investigated in the present study, especially some eastern European countries, are needed. Further, the present work proposes historical causes that may have contributed to shaping the current genetic variability of E. multilocularis in Europe.

Isolation of Toxoplasma gondii in cell culture: an alternative to bioassay.

Toxoplasma gondii is an apicomplexan protozoan parasite that can infect mammals and birds. The infection can cause acute toxoplasmosis and death in susceptible hosts. Bioassay using cats and mice has been the standard for the isolation of T. gondii from infected hosts for the past several decades. However, bioassay is labor-intensive, expensive, and involves using laboratory animals. To search alternative approaches and o work towards replacement of animal experiments, we summarized the key literature and conducted four experiments to isolate T. gondii in vitro by cell culture. A few heart tissue samples from animals with the highest antibody titers in a given collection were used for T. gondii isolation. These experiments included samples from five out of 51 wild ducks, four of 46 wild turkeys, six of 24 white-tailed deer, as well as from six kangaroos that had died with acute toxoplasmosis in a zoo. These experiments resulted in three isolates from five chronically infected wild ducks (60%), four isolates from four chronically infected wild turkeys (100%), one isolate from six chronically infected white-tailed deer (17%), and four isolates from six kangaroos with acute toxoplasmosis (67%). In addition, five isolates from the five chronically infected wild ducks were obtained by bioassay in mice, showing a 100% success rate, which is higher than the 60% rate by direct cell culture. These T. gondii isolates were successfully propagated in human foreskin fibroblast (HFF) or Vero cells, and genotyped by multilocus PCR-RFLP markers. The results showed that it is practical to isolate T. gondii directly in cell culture. Although the cell culture approach may not be as sensitive as the bioassay, it does provide an alternative that is simple, cost-effective, ethically more acceptable, and less time-sensitive to isolate T. gondii. In this paper we propose a procedure that may be applied and further optimized for isolation of T. gondii.

First cases of SARS-CoV-2 BA.2.86 in Denmark, 2023.

We describe 10 cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant BA.2.86 detected in Denmark, including molecular characteristics and results from wastewater surveillance that indicate that the variant is circulating in the country at a low level. This new variant with many spike gene mutations was classified as a variant under monitoring by the World Health Organization on 17 August 2023. Further global monitoring of COVID-19, BA.2.86 and other SARS-CoV-2 variants is highly warranted.

No Excess Cancer Risk among Veterinarians in Denmark, Finland, Iceland, Norway, and Sweden after the 1980s.

The cancer profile of veterinarians has received little research attention, despite the profession potentially being exposed to a wide range of known and suspected carcinogens. In this large-scale cohort study, we assessed cancer incidence in veterinarians in Denmark, Finland, Iceland, Norway, and Sweden, across more than 40 years (1961-2005). The cohort comprised 4708 veterinarians and 119,503 person-years at follow-up. The overall cancer incidence in veterinarians was close to the incidence in the total population in all countries and in all age groups. In male veterinarians, the standardized incidence ratios (SIR) in 1961-1990 were elevated for colon cancer (1.86, 95% confidence interval (CI) 1.39-2.44), prostate cancer (1.35, 95% CI 1.07-1.67), and especially skin melanoma (3.62, 95% CI 2.78-2.84), while there was no longer any statistically significant excess in the more recent follow-up period. Decreased SIRs were observed for lip cancer (0.11, 95% CI 0.00-0.62), laryngeal cancer (0.38, 95% CI 0.12-0.89), lung cancer (0.59, 95% CI 0.47-0.74), and stomach cancer (0.58, 95% CI 0.38-0.86), without a marked change in SIR over time. Non-significant excesses among male veterinarians were also observed in Hodgkin lymphoma (1961-1990 only), and leukaemia. This multi-country study indicates that there was an elevated incidence of several cancer types among male veterinarians before the 1990s but not after that. Some of the findings might rather be attributed to lifestyle factors and not directly to work conditions, but the excess risk of cancers of kidney and bladder, for example, might be related to work exposures.

Pulmonary cystic echinococcosis acquired during a short-term tourist travel.

Background: Cystic echinococcosis is non-endemic in Denmark and primarily diagnosed in migrants from endemic areas. Here, we report a case of pulmonary cystic echinococcosis in a Danish woman with no history of longer-term stays abroad, only holiday travelling to tourist destinations. This is the first case reported in international literature from Denmark where the causative parasite was identified to species and genotype level. Case: A 27-year-old pregnant Danish woman was admitted for examination because of haemoptysis for three months.Chest X-ray and computed tomography revealed a cystic structure in the left lung and a left-sided thoracotomy was performed to remove the cyst. Postoperative histopathological examination revealed a hyaline membrane and protoscoleces. Subsequently, infection with Echinococcus granulosus was confirmed by molecular methods. The causative agent was further characterised as E. granulosus sensu stricto G1, which is not known to have an established life cycle in Denmark. It was concluded that the infection was most likely acquired during a tourist travel to an endemic country. The patient was treated with albendazole for four weeks. Conclusion: This case of pulmonary cystic echinococcosis in a person who had lived in Denmark and had history of only short-term tourist travelling abroad highlights that the disease may be acquired during tourist travelling. Thus, a diagnosis of cystic echinococcosis should be considered not only in migrants from endemic countries but also in travellers upon incidental findings of a lung or liver cysts. The case also exemplifies the importance of reaching a diagnosis at species and genotype level.

Increase in invasive group A streptococcal infections and emergence of novel, rapidly expanding sub-lineage of the virulent Streptococcus pyogenes M1 clone, Denmark, 2023.

A highly virulent sub-lineage of the Streptococcus pyogenes M1 clone has been rapidly expanding throughout Denmark since late 2022 and now accounts for 30% of the new invasive group A streptococcal infections. We aimed to investigate whether a shift in variant composition can account for the high incidence rates observed over winter 2022/23, or if these are better explained by the impact of COVID-19-related restrictions on population immunity and carriage of group A Streptococcus.

A multi-country One Health foodborne outbreak simulation exercise: cross-sectoral cooperation, data sharing and communication.

Introduction: The awareness of scientists and policy makers regarding the requirement for an integrated One Health (OH) approach in responding to zoonoses has increased in recent years. However, there remains an overall inertia in relation to the implementation of practical cross-sector collaborations. Foodborne outbreaks of zoonotic diseases continue to affect the European population despite stringent regulations, evidencing the requirement for better 'prevent, detect and response' strategies. Response exercises play an essential role in the improvement of crisis management plans, providing the opportunity to test practical intervention methodologies in a controlled environment. Methods: The One Health European Joint Programme simulation exercise (OHEJP SimEx) aimed at practicing the OH capacity and interoperability across public health, animal health and food safety sectors in a challenging outbreak scenario. The OHEJP SimEx was delivered through a sequence of scripts covering the different stages of a Salmonella outbreak investigation at a national level, involving both the human food chain and the raw pet feed industry. Results: A total of 255 participants from 11 European countries (Belgium, Denmark, Estonia, Finland, France, Italy, Norway, Poland, Portugal, Sweden, the Netherlands) took part in national level two-day exercises during 2022. National evaluations identified common recommendations to countries aiming to improve their OH structure to establish formal communication channels between sectors, implement a common data sharing platform, harmonize laboratory procedures, and reinforce inter-laboratory networks within countries. The large proportion of participants (94%) indicated significant interest in pursuing a OH approach and desire to work more closely with other sectors. Discussion: The OHEJP SimEx outcomes will assist policy makers in implementing a harmonized approach to cross-sector health-related topics, by highlighting the benefits of cooperation, identifying gaps in the current strategies and suggesting actions required to better address foodborne outbreaks. Furthermore, we summarize recommendations for future OH simulation exercises, which are essential to continually test, challenge and improve national OH strategies.

Inactivation of Toxoplasma gondii in dry sausage and processed pork, and quantification of the pathogen in pig tissues prior to production.

Toxoplasma gondii is an important zoonotic foodborne parasite. Meat of infected animals appears to be a major source of infection in Europe. Pork is the most consumed meat in France, with dry sausages well represented. The risk of transmission via consumption of processed pork products is largely unknown, mainly since processing will affect viability but may not entirely inactivate all T. gondii parasites. We investigated the presence and concentration of T. gondii DNA in the shoulder, breast, ham, and heart of pigs orally inoculated with 1000 oocysts (n = 3) or tissue cysts (n = 3) and naturally infected pigs (n = 2), by means of magnetic capture qPCR (MC-qPCR). Muscle tissues of experimentally infected pigs were further used to evaluate the impact of manufacturing processes of dry sausages, including different concentrations of nitrates (0, 60, 120, 200 ppm), nitrites (0, 60, 120 ppm), and NaCl (0, 20, 26 g/kg), ripening (2 days at 16-24 °C) and drying (up to 30 days at 13 °C), by a combination of mouse bioassay, qPCR and MC-qPCR. DNA of T. gondii was detected in all eight pigs, including in 41.7% (10/24) of muscle samples (shoulder, breast and ham) and 87.5% (7/8) of hearts by MC-qPCR. The number of parasites per gram of tissue was estimated to be the lowest in the hams (arithmetic mean (M) = 1, standard deviation (SD) = 2) and the highest in the hearts (M = 147, SD = 233). However, the T. gondii burden estimates varied on the individual animal level, the tissue tested and the parasitic stage used for the experimental infection (oocysts or tissue cysts). Of dry sausages and processed pork, 94.4% (51/54) were positive for T. gondii by MC-qPCR or qPCR, with the mean T. gondii burden estimate equivalent to 31 parasites per gram (SD = 93). Only the untreated processed pork sample collected on the day of production was positive by mouse bioassay. The results suggest an uneven distribution of T. gondii in the tissues examined, and possibly an absence or a concentration below the detection limit in some of them. Moreover, the processing of dry sausages and processed pork with NaCl, nitrates, and nitrites has an impact on the viability of T. gondii from the first day of production. Results are valuable input for future risk assessments aiming to estimate the relative contribution of different sources of T. gondii human infections.

Identification of the tapeworm Mosgovoyia pectinata (Anoplocephalidae) in Faroese mountain hares (Lepus timidus).

The mountain hares (Lepus timidus L., 1758) in the Faroe Islands, an archipelago located in the North Atlantic, are known to be commonly infected by tapeworms, the identity of which was unknown. The mountain hare, which now populates 15 of the 18 islands, was introduced from Norway in 1855. In this study, tapeworms collected from four mountain hares from four geographic areas of the Faroe Islands were subjected to molecular identification using the nuclear ribosomal DNA (28S), the mitochondrial cytochrome oxidase subunit 1 (cox1) and the NADH dehydrogenase subunit 1 (nad1) genes. The results indicate unambiguously that the tapeworms were Mosgovoyia pectinata (Goeze, 1782) (Cestoda: Anoplocephalidae sensu stricto). The phylogenetic position and origin of the Faroese M. pectinata are discussed. Given that the parasite is quite common in Norway, from where the mountain hares were introduced, it is conceivable that co-introduction of M. pectinata from Norway to the Faroe Islands took place. The phylogenetic analyses revealed high similarity of the M. pectinata sequences from three regions and the position of the Faroese isolate as the sister lineage of the isolates from Finland and East Siberia.