No Substantial Histopathologic Changes in Mops condylurus Bats Naturally Infected with Bombali Virus, Kenya.

We found similar mild perivascular inflammation in lungs of Bombali virus-positive and -negative Mops condylurus bats in Kenya, indicating the virus is well-tolerated. Our findings indicate M. condylurus bats may be a reservoir host for Bombali virus. Increased surveillance of these bats will be important to reduce potential virus spread.

Stable Levels of Antibodies Against Unrelated Toxoid Vaccines After COVID-19: COVID-19 Infection Does Not Affect Toxoid Vaccine Antibody Levels.

Background: Lymphopenia is common in COVID-19. This has raised concerns that COVID-19 could affect the immune system akin to measles infection, which causes immune amnesia and a reduction in protective antibodies. Methods: We recruited COVID-19 patients (n = 59) in Helsinki, Finland, and collected plasma samples on 2 to 3 occasions during and after infection. We measured IgG antibodies to diphtheria toxin, tetanus toxoid, and pertussis toxin, along with total IgG, SARS-CoV-2 spike protein IgG, and neutralizing antibodies. We also surveyed the participants for up to 17 months for long-term impaired olfaction as a proxy for prolonged post-acute COVID-19 symptoms. Results: No significant differences were found in the unrelated vaccine responses while the serological response against COVID-19 was appropriate. During the acute phase of the disease, the SARSCoV-2 IgG levels were lower in outpatients when compared to inpatients. SARS-CoV-2 serology kinetics matched expectations. In the acute phase, anti-tetanus and anti-diphtheria IgG levels were lower in patients with prolonged impaired olfaction during follow up than in those without. Conclusions: We could not detect significant decline in overall humoral immunity during or after COVID-19 infection. In severe COVID-19, there appears to be a temporary decline in total IgG levels.

SARS-CoV-2 indoor environment contamination with epidemiological and experimental investigations.

SARS-CoV-2 has been detected both in air and on surfaces, but questions remain about the patient-specific and environmental factors affecting virus transmission. Additionally, more detailed information on viral sampling of the air is needed. This prospective cohort study (N = 56) presents results from 258 air and 252 surface samples from the surroundings of 23 hospitalized and eight home-treated COVID-19 index patients between July 2020 and March 2021 and compares the results between the measured environments and patient factors. Additionally, epidemiological and experimental investigations were performed. The proportions of qRT-PCR-positive air (10.7% hospital/17.6% homes) and surface samples (8.8%/12.9%) showed statistical similarity in hospital and homes. Significant SARS-CoV-2 air contamination was observed in a large (655.25 m3 ) mechanically ventilated (1.67 air changes per hour, 32.4-421 L/s/patient) patient hall even with only two patients present. All positive air samples were obtained in the absence of aerosol-generating procedures. In four cases, positive environmental samples were detected after the patients had developed a neutralizing IgG response. SARS-CoV-2 RNA was detected in the following particle sizes: 0.65-4.7 µm, 7.0-12.0 µm, >10 µm, and <100 µm. Appropriate infection control against airborne and surface transmission routes is needed in both environments, even after antibody production has begun.

Experimental Infection of Mink with SARS-COV-2 Omicron Variant and Subsequent Clinical Disease.

We report an experimental infection of American mink with SARS-CoV-2 Omicron variant and show that mink remain positive for viral RNA for days, experience clinical signs and histopathologic changes, and transmit the virus to uninfected recipients. Preparedness is crucial to avoid spread among mink and spillover to human populations.

Copper-Silver Nanohybrids: SARS-CoV-2 Inhibitory Surfaces.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains a severe health threat. The COVID-19 infections occurring in humans and animals render human-animal interfaces hot spots for spreading the pandemic. Lessons from the past point towards the antiviral properties of copper formulations; however, data showing the "contact-time limit" surface inhibitory efficacy of copper formulations to contain SARS-CoV-2 are limited. Here, we show the rapid inhibition of SARS-CoV-2 after only 1 and 5 min on two different surfaces containing copper-silver (Cu-Ag) nanohybrids. We characterized the nanohybrids' powder and surfaces using a series of sophisticated microscopy tools, including transmission and scanning electron microscopes (TEM and SEM) and energy-dispersive X-ray spectroscopy (EDX). We used culturing methods to demonstrate that Cu-Ag nanohybrids with high amounts of Cu (~65 and 78 wt%) and lower amounts of Ag (~7 and 9 wt%) inhibited SARS-CoV-2 efficiently. Collectively, the present work reveals the rapid SARS-CoV-2 surface inhibition and the promising application of such surfaces to break the SARS-CoV-2 transmission chain. For example, such applications could be invaluable within a hospital or live-stock settings, or any public place with surfaces that people frequently touch (i.e., public transportation, shopping malls, elevators, and door handles) after the precise control of different parameters and toxicity evaluations.

Severe Acute Respiratory Syndrome Coronavirus 2 in Farmed Mink (Neovison vison), Poland.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiologic agent of coronavirus disease and has been spreading worldwide since December 2019. The virus can infect different animal species under experimental conditions, and mink on fur farms in Europe and other areas are susceptible to SARS-CoV-2 infection. We investigated SARS-CoV-2 infection in 91 mink from a farm in northern Poland. Using reverse transcription PCR, antigen detection, and next-generation sequencing, we confirmed that 15 animals were positive for SARS-CoV-2. We verified this finding by sequencing full viral genomes and confirmed a virus variant that has sporadic mutations through the full genome sequence in the spike protein (G75V and C1247F). We were unable to find other SARS-CoV-2 sequences simultaneously containing these 2 mutations. Country-scale monitoring by veterinary inspection should be implemented to detect SARS-CoV-2 in other mink farms.

Comparative Genomics of 42 Arcanobacterium phocae Strains.

For the last 13 years, the fur industry in Europe has suffered from epidemic spouts of a severe necrotizing pyoderma. It affects all species currently farmed for fur and causes animal welfare problems and significant losses to the farmers. The causative agent of this disease was identified as Arcanobacterium phocae. Previously, this bacterium has been isolated from seals and other marine mammals, apparently causing wound and lung infections. Attempts at antibiotic treatment have been unsuccessful and the current advice on preventing the disease is to cull all animals with clinical signs. This poses an urgent question regarding possible vaccine development, as well as the need for further understanding of the pathogenicity of this organism. This study compared the whole genomes of 42 A. phocae strains isolated from seals, blue foxes, finnraccoons, mink and otter. The sequences were created using the Illumina technology and annotations were done using the RAST pipeline. A phylogenetic analysis identified a clear separation between the seal strains and the fur-animal-derived isolates, but also indicated that the bacterium readily adapts to new environments and host species with reasonable diversity. A pan- and core-genome was created and analyzed for proteins. A further analysis identified several virulence factors as well as multiple putative and secreted proteins of special interest for vaccine development.

Kinetics of Neutralizing Antibodies of COVID-19 Patients Tested Using Clinical D614G, B.1.1.7, and B 1.351 Isolates in Microneutralization Assays.

Increasing evidence suggests that some newly emerged SARS-CoV-2 variants of concern (VoCs) resist neutralization by antibodies elicited by the early-pandemic wild-type virus. We applied neutralization tests to paired recoveree sera (n = 38) using clinical isolates representing the first wave (D614G), VoC1, and VoC2 lineages (B.1.1.7 and B 1.351). Neutralizing antibodies inhibited contemporary and VoC1 lineages, whereas inhibition of VoC2 was reduced 8-fold, with 50% of sera failing to show neutralization. These results provide evidence for the increased potential of VoC2 to reinfect previously SARS-CoV-infected individuals. The kinetics of NAbs in different patients showed similar decline against all variants, with generally low initial anti-B.1.351 responses becoming undetectable, but with anti-B.1.1.7 NAbs remaining detectable (>20) for months after acute infection.

Survival of SARS-CoV-2 on Clothing Materials.

In order to plan and execute proper preventative measures against COVID-19, we need to understand how SARS-CoV-2 is transmitted. It has been shown to remain infectious on surfaces from hours to days depending on surface type and environmental factors. The possibility of transmission through fur animals and contaminated pelts, along with the safety of those working with them, is a major concern. SARS-CoV-2 can infect minks and raccoon dogs and has spread to mink farms in numerous countries. Here, we studied the stability of SARS-CoV-2 on blue fox, Finn raccoon, and American mink pelt, fake fur, cotton, plastic, faux leather, and polyester and tested its inactivation by UV light and heat treatment. We detected infectious virus up to 5 days on plastic, up to 1 day on fake fur, less than a day on cotton, polyester, and faux leather, and even 10 days on mink fur. UV light failed to inactivate SARS-CoV-2 on pelts, most likely due to the mechanical protection by the fur. Hence, it should not be used to inactivate the virus on fur products, and its use for other surfaces should also be considered carefully. Heat treatment at 60°C for 1 h inactivated the virus on all surfaces and is a promising method to be applied in practice. This study helps prevent further spread of COVID-19 by increasing our understanding about risks of SARS-CoV-2 spread through contaminated clothing materials and giving important information needed to improve safety of those working in the production line as well as people using the products.

Range Expansion of Bombali Virus in Mops condylurus Bats, Kenya, 2019.

Previously identified only in Sierra Leone, Guinea, and southeastern Kenya, Bombali virus-infected Mops condylurus bats were recently found ¼750 km away in western Kenya. This finding supports the role of M. condylurus bats as hosts and the potential for Bombali virus circulation across the bats' range in sub-Saharan Africa.

Comparison of Streptococcus halichoeri isolates from canine and fur animal infections: biochemical patterns, molecular characteristics and genetic relatedness.

BACKGROUND: Streptococcus halichoeri infections have been reported in grey seals, a European badger, a Stellar sea lion and humans, but its presence in companion and fur animals is unknown. Since 2010, S. halichoeri-like bacteria (SHL) have been isolated from fur animals and dogs in Finland. Our aim was to retrospectively investigate laboratory records for SHL from canine and fur animal infections, characterize the isolates and compare their genetic relatedness in relation to three reference strains: CCUG 48324T, originating from a grey seal, and strains 67100 and 61265, originally isolated from humans. RESULTS: A total of 138 and 36 SHLs from canine and fur animal infections, respectively, were identified in the laboratory records. SHL was commonly associated with skin infections, but rarely as the only species. A set of 49 canine and 23 fur animal SHLs were further characterized. MALDI-TOF confirmed them as being S. halichoeri. The growth characteristics were consistent with the original findings, but isolates were catalase positive. In total, 17 distinct API 20 Strep patterns were recorded among all 75 isolates tested, of which pattern 5563100 was the most common (n = 30). Antimicrobial resistance to erythromycin and clindamycin was common in canine isolates, but rare in fur animal isolates. Three clusters were observed by PFGE, and 16S rRNA sequencing revealed 98.1-100% similarities with the human strains and 98.1-99.5% with the seal strain. A phylogenetic tree of concatenated 16S rRNA and rpoB revealed closely related isolates with two clades. Fifteen canine isolates were identical to the human strains based on concatenated 16S rRNA and rpoB sequencing. CONCLUSIONS: Streptococcus halichoeri appears to be quite a common bacterial species in the skin of dogs and fur animals. The clinical significance of S. halichoeri is uncertain, as it was rarely isolated as a monoculture. No apparent temporal or spatial clustering was detected, but isolates from different sources were genetically very similar. Because many canine isolates were genetically similar to the human reference strains, transmission between dogs and humans may be possible. WGS sequencing of strains from different sources is needed to further investigate the epidemiology and virulence of S. halichoeri.

Orthohantavirus Isolated in Reservoir Host Cells Displays Minimal Genetic Changes and Retains Wild-Type Infection Properties.

: Orthohantaviruses are globally emerging zoonotic pathogens. While the reservoir host role of several rodent species is well-established, detailed research on the mechanisms of host-othohantavirus interactions has been constrained by the lack of an experimental system that is able to effectively replicate natural infections in controlled settings. Here we report the isolation, and genetic and phenotypic characterization of a novel Puumala orthohantavirus (PUUV) in cells derived from its reservoir host, the bank vole. The isolation process resulted in cell culture infection that evaded antiviral responses, persisted cell passaging, and had minor viral genome alterations. Critically, experimental infections of bank voles with the new isolate resembled natural infections in terms of viral load and host cell distribution. When compared to an attenuated Vero E6 cell-adapted PUUV Kazan strain, the novel isolate demonstrated delayed virus-specific humoral responses. A lack of virus-specific antibodies was also observed during experimental infections with wild-type PUUV, suggesting that delayed seroconversion could be a general phenomenon during orthohantavirus infection in reservoir hosts. Our results demonstrate that orthohantavirus isolation on cells derived from a vole reservoir host retains wild-type infection properties and should be considered the method of choice for experimental infection models to replicate natural processes.

Streptococcus halichoeri: Comparative Genomics of an Emerging Pathogen.

Streptococcus halichoeri is an emerging pathogen with a variety of host species and zoonotic potential. It has been isolated from grey seals and other marine mammals as well as from human infections. Beginning in 2010, two concurrent epidemics were identified in Finland, in fur animals and domestic dogs, respectively. The fur animals suffered from a new disease fur animal epidemic necrotic pyoderma (FENP) and the dogs presented with ear infections with poor treatment response. S. halichoeri was isolated in both studies, albeit among other pathogens, indicating a possible role in the disease etiologies. The aim was to find a possible common origin of the fur animal and dog isolates and study the virulence factors to assess pathogenic potential. Isolates from seal, human, dogs, and fur animals were obtained for comparison. The whole genomes were sequenced from 20 different strains using the Illumina MiSeq platform and annotated using an automatic annotation pipeline RAST. The core and pangenomes were formed by comparing the genomes against each other in an all-against-all comparison. A phylogenetic tree was constructed using the genes of the core genome. Virulence factors were assessed using the Virulence Factor Database (VFDB) concentrating on the previously confirmed streptococcal factors. A core genome was formed which encompassed approximately half of the genes in Streptococcus halichoeri. The resulting core was nearly saturated and would not change significantly by adding more genomes. The remaining genes formed the pangenome which was highly variable and would still evolve after additional genomes. The results highlight the great adaptability of this bacterium possibly explaining the ease at which it switches hosts and environments. Virulence factors were also analyzed and were found primarily in the core genome. They represented many classes and functions, but the largest single category was adhesins which again supports the marine origin of this species.

Development and validation of nucleic acid tests to diagnose Aleutian mink disease virus.

Aleutian disease (AD), caused by Aleutian mink disease virus (AMDV), causes significant welfare problems to mink, and financial losses to the farmers. As there is no vaccine or treatment available, reliable diagnostics is important for disease control. Here, we set up a probe-based real-time PCR (NS1-probe-PCR) to detect all strains of AMDV. PCR was validated and compared to two other real-time PCR methods (pan-AMDV- and pan-AMDO-PCR) currently used for AMDV diagnostics in Finland. The NS1-probe-PCR had a similar detection limit of 20 copies/reaction based on plasmid dilution series, and similar or better diagnostic sensitivity, when evaluated using spleen samples from mink, and stool samples from mink and foxes. None of the three PCR tests cross-reacted with other parvoviruses. The NS1-probe-PCR also showed a significantly higher specificity than the pan-AMDO-PCR with spleen samples and the best specificity with stool samples. Furthermore, it produced the results more rapidly than the other two PCRs making it a promising tool for both diagnostic and research purposes.

Co-circulation of highly diverse Aleutian mink disease virus strains in Finland.

Aleutian mink disease virus (AMDV) is the causative agent of Aleutian disease (AD), which affects mink of all genotypes and also infects other mustelids such as ferrets, martens and badgers. Previous studies have investigated diversity in Finnish AMDV strains, but these studies have been restricted to small parts of the virus genome, and mostly from newly infected farms and free-ranging mustelids. Here, we investigated the diversity and evolution of Finnish AMDV strains by sequencing the complete coding sequences of 31 strains from mink originating from farms differing in their virus history, as well as from free-ranging mink. The data set was supplemented with partial genomes obtained from 26 strains. The sequences demonstrate that the Finnish AMDV strains have considerable diversity, and that the virus has been introduced to Finland in multiple events. Frequent recombination events were observed, as well as variation in the evolutionary rate in different parts of the genome and between different branches of the phylogenetic tree. Mink in the wild carry viruses with high intra-host diversity and are occasionally even co-infected by two different strains, suggesting that free-ranging mink tolerate chronic infections for extended periods of time. These findings highlight the need for further sampling to understand the mechanisms playing a role in the evolution and pathogenesis of AMDV.

Family history influences the tumor characteristics and prognosis of breast cancers developing during postmenopausal hormone therapy.

Long term use of postmenopausal hormone therapy (HT) has been reported to increase breast cancer risk. On the other hand, observational studies suggest that breast cancers diagnosed during HT may have a more favorable prognosis. While family history is a risk factor for breast cancer, and genetic factors also influence prognosis, the role of family history in combination with HT use has been little studied. We investigated the relationship between HT, family history, and prognosis in 584 (267 exposed) familial and 952 (460 exposed) non-familial breast cancer cases, using three survival end points: death from breast cancer (BCS), distant disease free survival (DDFS), and local recurrence free survival (LRFS). Among non-familial cases, HT was associated with better BCS (HR 0.63, 95% CI 0.41-0.94; p = 0.025), and DDFS (HR 0.58, 95% CI 0.40-0.85; p = 0.005), with a consistent but not statistically significant effect in LRFS. This effect was not seen in familial cases (HR > 1.0), and family history was found to interact with HT in BCS (p(interaction) = 0.0067) (BC-death) and DDFS (p(interaction) = 0.0070). There was phenotypic heterogeneity between HT-associated tumors in familial and non-familial cases, particularly on estrogen receptor (ER) status, although the interaction between HT and family history appears to be at least partially independent of these markers (p = 0.0370 after adjustment for standard prognostic factors). If confirmed by further studies, our results suggest that family history should be taken into consideration in clinical counseling before beginning a HT regimen.

Experimental Infection of Mink Enforces the Role of Arcanobacterium phocae as Causative Agent of Fur Animal Epidemic Necrotic Pyoderma (FENP).

Fur Animal Epidemic Necrotic Pyoderma (FENP) is a severe, often lethal infectious disease affecting all three fur animal species: mink (Neovision vision), foxes (Vulpes lagopus) and finnraccoons (Nyctereutes procyonoides). Previous studies showed an association between Arcanobacterium phocae and FENP. An experimental infection was conducted to confirm the ability of A. phocae to infect mink either alone or concurrently with a novel Streptococcus sp. found together with A. phocae in many cases of FENP. Different inoculation methods were tested to study possible routes of transmission. Typical signs, and gross- and histopathological findings for FENP were detected when naïve mink were infected with the tissue extract of mink with FENP, using a subcutaneous/ intradermal infection route. Edema, hemorrhage, necrosis and pus formation were detected in the infection site. A pure culture preparation of A. phocae alone or concurrently with the novel Streptococcus sp. caused severe acute signs of lethargy, apathy and anorexia and even mortality. The histopathological findings were similar to those found in naturally occurring cases of FENP. In contrast, the perorally infected mink presented no clinical signs nor any gross- or histopathological lesions. This study showed that A. phocae is able to cause FENP. The study also indicated that predisposing factors such as the environment, the general condition of the animals, temperature and skin trauma contribute to the development of the disease.

GEOGRAPHIC DISTRIBUTION AND MOLECULAR DIVERSITY OF BARTONELLA SPP. INFECTIONS IN MOOSE (ALCES ALCES) IN FINLAND.

Moose, Alces alces (Artiodactyla: Cervidae) in Finland are heavily infested with deer keds, Lipoptena cervi (Diptera: Hippoboschidae). The deer ked, which carries species of the genus Bartonella, has been proposed as a vector for the transmission of bartonellae to animals and humans. Previously, bartonella DNA was found in deer keds as well as in moose blood collected in Finland. We investigated the prevalence and molecular diversity of Bartonella spp. infection from blood samples collected from free-ranging moose. Given that the deer ked is not present in northernmost Finland, we also investigated whether there were geographic differences in the prevalence of bartonella infection in moose. The overall prevalence of bartonella infection was 72.9% (108/148). Geographically, the prevalence was highest in the south (90.6%) and lowest in the north (55.9%). At least two species of bartonellae were identified by multilocus sequence analysis. Based on logistic regression analysis, there was no significant association between bartonella infection and either age or sex; however, moose from outside the deer ked zone were significantly less likely to be infected (P<0.015) than were moose hunted within the deer ked zone.

Development and evaluation of a real-time EBOV-L-RT-qPCR for detection of Zaire ebolavirus.

An RT-qPCR targeting EBOV-L including the preceding RNA extraction protocol were set up and evaluated.

Characterization of a new epidemic necrotic pyoderma in fur animals and its association with Arcanobacterium phocae infection.

A new type of pyoderma was detected in Finnish fur animals in 2007. The disease continues to spread within and between farms, with severe and potentially fatal symptoms. It compromises animal welfare and causes considerable economic losses to farmers. A case-control study was performed in 2010-2011 to describe the entity and to identify the causative agent. Altogether 99 fur animals were necropsied followed by pathological and microbiological examination. The data indicated that the disease clinically manifests in mink (Neovison vison) by necrotic dermatitis of the feet and facial skin. In finnraccoons (Nyctereutes procyonoides), it causes painful abscesses in the paws. Foxes (Vulpes lagopus) are affected by severe conjunctivitis and the infection rapidly spreads to the eyelids and facial skin. A common finding at necropsy was necrotic pyoderma. Microbiological analysis revealed the presence of a number of potential causative agents, including a novel Streptococcus sp. The common finding from all diseased animals of all species was Arcanobacterium phocae. This bacterium has previously been isolated from marine mammals with skin lesions but this is the first report of A. phocae isolated in fur animals with pyoderma. The results obtained from this study implicate A. phocae as a potential causative pathogen of fur animal epidemic necrotic pyoderma (FENP) and support observations that the epidemic may have originated in a species-shift of the causative agent from marine mammals. The variable disease pattern and the presence of other infectious agents (in particular the novel Streptococcus sp.) suggest a multifactorial etiology for FENP, and further studies are needed to determine the environmental, immunological and infectious factors contributing to the disease.