Longitudinal positron emission tomography and postmortem analysis reveals widespread neuroinflammation in SARS-CoV-2 infected rhesus macaques.

BACKGROUND: Coronavirus disease 2019 (COVID-19) patients initially develop respiratory symptoms, but they may also suffer from neurological symptoms. People with long-lasting effects after acute infections with severe respiratory syndrome coronavirus 2 (SARS-CoV-2), i.e., post-COVID syndrome or long COVID, may experience a variety of neurological manifestations. Although we do not fully understand how SARS-CoV-2 affects the brain, neuroinflammation likely plays a role. METHODS: To investigate neuroinflammatory processes longitudinally after SARS-CoV-2 infection, four experimentally SARS-CoV-2 infected rhesus macaques were monitored for 7 weeks with 18-kDa translocator protein (TSPO) positron emission tomography (PET) using [18F]DPA714, together with computed tomography (CT). The baseline scan was compared to weekly PET-CTs obtained post-infection (pi). Brain tissue was collected following euthanasia (50 days pi) to correlate the PET signal with TSPO expression, and glial and endothelial cell markers. Expression of these markers was compared to brain tissue from uninfected animals of comparable age, allowing the examination of the contribution of these cells to the neuroinflammatory response following SARS-CoV-2 infection. RESULTS: TSPO PET revealed an increased tracer uptake throughout the brain of all infected animals already from the first scan obtained post-infection (day 2), which increased to approximately twofold until day 30 pi. Postmortem immunohistochemical analysis of the hippocampus and pons showed TSPO expression in cells expressing ionized calcium-binding adaptor molecule 1 (IBA1), glial fibrillary acidic protein (GFAP), and collagen IV. In the hippocampus of SARS-CoV-2 infected animals the TSPO+ area and number of TSPO+ cells were significantly increased compared to control animals. This increase was not cell type specific, since both the number of IBA1+TSPO+ and GFAP+TSPO+ cells was increased, as well as the TSPO+ area within collagen IV+ blood vessels. CONCLUSIONS: This study manifests [18F]DPA714 as a powerful radiotracer to visualize SARS-CoV-2 induced neuroinflammation. The increased uptake of [18F]DPA714 over time implies an active neuroinflammatory response following SARS-CoV-2 infection. This inflammatory signal coincides with an increased number of TSPO expressing cells, including glial and endothelial cells, suggesting neuroinflammation and vascular dysregulation. These results demonstrate the long-term neuroinflammatory response following a mild SARS-CoV-2 infection, which potentially precedes long-lasting neurological symptoms.

Tolerant and intolerant macaques show different levels of structural complexity in their vocal communication.

We tested the social complexity hypothesis which posits that animals living in complex social environments should use complex communication systems. We focused on two components of vocal complexity: diversity (number of categories of calls) and flexibility (degree of gradation between categories of calls). We compared the acoustic structure of vocal signals in groups of macaques belonging to four species with varying levels of uncertainty (i.e. complexity) in social tolerance (the higher the degree of tolerance, the higher the degree of uncertainty): two intolerant species, Japanese and rhesus macaques, and two tolerant species, Tonkean and crested macaques. We recorded the vocalizations emitted by adult females in affiliative, agonistic and neutral contexts. We analysed several acoustic variables: call duration, entropy, time and frequency energy quantiles. The results showed that tolerant macaques displayed higher levels of vocal diversity and flexibility than intolerant macaques in situations with a greater number of options and consequences, i.e. in agonistic and affiliative contexts. We found no significant differences between tolerant and intolerant macaques in the neutral context where individuals are not directly involved in social interaction. This shows that species experiencing more uncertain social interactions displayed greater vocal diversity and flexibility, which supports the social complexity hypothesis.

Determining overweight and underweight with a new weight-for-height index in captive group-housed macaques.

Housing primates in naturalistic groups provides social benefits relative to solitary housing. However, food intake may vary across individuals, possibly resulting in overweight and underweight individuals. Information on relative adiposity (the amount of fat tissue relative to body weight) is needed to monitor overweight and underweight of group-housed individuals. However, the upper and lower relative adiposity boundaries are currently only known for macaques living solitarily in small cages. We determined the best measure of relative adiposity and explored the boundaries of overweight and underweight to investigate their incidence in group-housed adult male and female rhesus macaques and long-tailed macaques living in spacious enclosures at the Biomedical Primate Research Centre (BPRC), the Netherlands. During yearly health checks different relative adiposity measures were obtained. For long-tailed macaques, comparable data on founder and wild animals were also available. Weight-for-height indices (WHI) with height to the power of 3.0 (WHI3.0) for rhesus macaques and 2.7 (WHI2.7) for long-tailed macaques were optimally independent of height and were highly correlated with other relative adiposity measures. The boundary for overweight was similar in group-housed and solitary-housed macaques. A lower boundary for underweight, based on 2% body fat similar to wild primates, gave a better estimate for underweight in group-housed macaques. We propose that for captive group-housed rhesus macaques relative adiposity should range between 42 and 67 (WHI3.0) and for long-tailed macaques between 39 and 62 (WHI2.7). The majority of group-housed macaques in this facility have a normal relative adiposity, a considerable proportion (17-23%) is overweight, and a few (0-3%) are underweight.

25-OH-vitamin D, parathyroid hormone, and calcium serum levels in captive common marmosets (Callithrix jacchus): Reference values and effect of age, sex, season, and closure of long bone epiphyses.

BACKGROUND: To date, reference values for 25-OH-vitamin D, parathyroid hormone (PTH), and calcium in serum of common marmosets (Callithrix jacchus) based on a large sample size are not available. METHODS: Serum reference values for these parameters were determined and correlated with sex, age, season of sampling, and time of long bone epiphyseal closure in captive-housed marmosets. RESULTS AND CONCLUSIONS: The 90% reference range for serum 25-OH-vitamin D is 47.40-370.4 nmol/L, for PTH 2.10-30.51 pmol/L, and for calcium 2.08-2.63 mmol/L. Lower levels of vitamin D were measured in fall compared with the other seasons. Levels of PTH were higher in males than in females, and calcium levels were lower in younger animals compared with older marmosets. No other effects of age, sex, season, or timing of growth plate closure were found.

Defining predictive factors for reproductive output in captive common marmosets (Callithrix jacchus).

Common marmosets (Callithrix jacchus) demonstrate variations in reproductive output, not only in terms of total reproductive output during a lifetime but also in litter size per parturition. The present study explores factors, such as parents' litter size, parturition number, maternal body weight at conception and maternal age, which may account for this variation. A retrospective analysis of clinical records of a captive breeding colony was conducted over a 9-year period yielding reproductive summaries of 26 dams and 22 sires producing a total of 115 litters. Dams born from litters of ≤2 (N = 20) more often produced litters of ≤2, whereas dams born from litters of >2 (N = 6) more often produced litters of >2 (p < 0.05). The dams' maternal body weight at the time of conception had also a significant effect on subsequent litter size. In addition, the chance of triplets was higher after the second parturition. Maternal age, interbirth interval, and season of birth had no effect on litter size. Factors relating to the sire had a negligible effect on the size of the litter. Multivariate statistical modeling revealed that the dams' original litter size, maternal bodyweight at conception and parturition number are determining factors for the number of babies per litter. This study identified factors determining marmoset litter size, some of which (maternal litter size) are novel to this study and were not reported previously. Further exploration of the potential role of maternal litter size as a determinant of the litter sizes produced by marmoset breeders is warranted.

Comparative virulence of in vitro-cultured primate- and pig-associated Helicobacter suis strains in a BALB/c mouse and a Mongolian gerbil model.

BACKGROUND: Helicobacter suis (H. suis) is the most prevalent gastric non-H. pylori Helicobacter species in humans. This bacterium mainly colonizes the stomach of pigs, but it has also been detected in the stomach of nonhuman primates. The aim of this study was to obtain better insights into potential differences between pig- and primate-associated H. suis strains in virulence and pathogenesis. MATERIALS AND METHODS: In vitro-isolated H. suis strains obtained from pigs, cynomolgus monkeys (Macaca fascicularis), and rhesus monkeys (Macaca mulatta) were used for intragastric inoculation of BALB/c mice and Mongolian gerbils. Nine weeks and six months later, samples of the stomach of inoculated and control animals were taken for PCR analysis and histopathological examination. RESULTS: The cynomolgus monkey-associated H. suis strain only colonized the stomach of mice, but not of Mongolian gerbils. All other H. suis strains colonized the stomach in both rodent models. In all colonized animals, severe gastric inflammation was induced. Gastric lymphoid follicles and destruction of the antral epithelium were observed in infected gerbils, but not in mice. Infection with both pig- and primate-associated H. suis strains evoked a similar marked Th17 response in mice and gerbils, accompanied by increased CXCL-13 expression levels. CONCLUSIONS: Apart from the cynomolgus monkey-associated strain which was unable of colonizing the stomach of Mongolian gerbils, no substantial differences in virulence were found in rodent models between in vitro-cultured pig-associated, cynomolgus monkey-associated and rhesus monkey-associated H. suis strains. The experimental host determines the outcome of the immune response against H. suis infection, rather than the original host.

Peptococcus simiae sp. nov., isolated from rhesus macaque faeces and emended description of the genus Peptococcus.

A study of the faecal microbiome in three healthy female rhesus macaques revealed the presence of a novel obligately anaerobic, chemoorganoheterotrophic, non-sporing, coccoid, non-motile, Gram-stain-positive bacterial species. Three strains of this species, designated as M108T, M916-1/1, and M919-2/1, were non-haemolytic, H2S-positive, catalase-positive, bile- and NaCl-sensitive and required peptone for growth. Strains also were asaccharolytic, able to utilize sulfite, thiosulfate and elemental sulfur as electron acceptors, and produced acetic and butyric acids as metabolic end-products. Strain M108T is characterized by the prevalence of C14 : 0, C16 : 0 and C18 : 1ω9cis dimethyl acetal among the cellular fatty acids, and the presence of MK-10 menaquinone. The DNA G+C content was found to be 51 mol%. Phylogenetic analysis of partial 16S rRNA gene sequences of strains M108T, M916-1/1 and M919-2/1 placed these strains into the genus Peptococcus (family Peptococcaceae). On the basis of phenotypic and genotypic properties we conclude that these strains represent a novel bacterial species for which the name Peptococcus simiae sp. nov. is proposed. The type strain is M108T (=DSM 100347T=VKM B-2932T).

Acute-phase responses in healthy and diseased rhesus macaques (Macaca mulatta).

Five acute-phase reactants-serum amyloid A (SAA), C-reactive protein (CRP), haptoglobin, albumin, and iron-were measured using commercially available assays in 110 healthy rhesus macaques (Macaca mulatta), and reference intervals were established for future use in health monitoring of this species. Reference intervals established were as follows: SAA, 29.5-87.7 mg/L; CRP, 0-17.5 mg/L; haptoglobin, 354.3-2,414.7 mg/ L; albumin, 36.1-53.0 g/L; and iron, 13.3-40.2 micromol/L. Furthermore, changes in the acute-phase reactants were studied in two additional groups of animals: eight rhesus macaques suffering from acute traumatic injuries and nine rhesus macaques experimentally infected with Mycobacterium tuberculosis reflecting a chronic active inflammation. In animals with inflammation, SAA and haptoglobin concentrations were moderately increased, while CRP increased more than 200-fold. In addition, marked decreases in albumin and iron concentrations were observed. These results show that SAA, CRP, and haptoglobin are positive acute-phase proteins, whereas albumin and iron are negative acute-phase reactants in rhesus macaques.

Hematological and Serum Biochemical Reference Intervals for Alphaxalone Sedated Common Marmosets (Callithrix jacchus).

Marmosets are routinely used in biomedical research, therefore there is an increasing need for updated reference intervals calculated using a large sample size, correct statistics, and considering different variables. Hematological and biochemical values from 472 healthy common marmosets sedated with alphaxalone were collected over a ten-year period (2013-2023). The variables assumed to have influenced the blood-based parameters were compared, i.e., sex, age, housing condition, pregnancy, and contraceptive use. Reference intervals were calculated based on observed percentiles without parametric assumptions, and with parametric assumptions following Box-Cox transformation. Juvenile marmosets showed increased ALP, phosphate, WBC, lymphocyte count, and basophil count and decreased levels of GGT and Fe compared to adults. Marmosets housed strictly indoors showed increased ALT and GGT levels and decreased levels of total bilirubin and neutrophil count compared to marmosets housed with outdoor access. Pregnant marmosets showed increased ALP, total bilirubin, neutrophil count, monocyte count, and basophil count, and decreased levels of AST, ALT, cholesterol, Fe, and lymphocyte count compared to non-pregnant marmosets. Etonogestrel contracepted marmosets showed decreased P-LCR compared to females who were not contracepted. Updated reference intervals will aid researchers and veterinarians in identifying physiological and pathological changes, as well as improve the reproducibility of research in this species.

Comparison of three different sedative-anaesthetic protocols (ketamine, ketamine-medetomidine and alphaxalone) in common marmosets (Callithrix jacchus).

BACKGROUND: Handling of common marmoset (Callithrix jacchus) usually requires chemical restraint. Ketamine has been associated with muscle damage in primates, while common marmosets, compared to other primates, additionally display an exceptional high sensitivity to ketamine-associated side-effects. Notably, muscle twitching movements of limbs and hands, and a marked increase in salivation are observed. We investigated two alternative intramuscular (i.m.) immobilisation protocols against ketamine (50 mg/kg; protocol 1) in a double-blind randomised crossover study in ten healthy adult common marmosets for use as a safe reliable, short-term immobilisation and sedation. These protocols comprised: alphaxalone (12 mg/kg; protocol 2) and 25 mg/kg ketamine combined with 0.50 mg/kg medetomidine (reversal with 2.5 mg/kg atipamezole; protocol 3A). Following completion and unblinding, the project was extended with an additional protocol (3B), comprising 25 mg/kg ketamine combined with 0.05 mg/kg medetomidine (reversal with 0.25 mg/kg atipamezole, twice with 35 min interval). RESULTS: All protocols in this study provided rapid onset (induction times <5 min) of immobilisation and sedation. Duration of immobilisation was 31.23 ± 22.39 min, 53.72 ± 13.08 min, 19.73 ± 5.74 min, and 22.78 ± 22.37 min for protocol 1, 2, 3A, and 3B, respectively. Recovery times were 135.84 ± 39.19 min, 55.79 ± 11.02 min, 405.46 ± 29.81 min, and 291.91 ± 80.34 min, respectively. Regarding the quality, and reliability (judged by pedal withdrawal reflex, palpebral reflex and muscle tension) of all protocols, protocol 2 was the most optimal. Monitored vital parameters were within clinically acceptable limits during all protocols and there were no fatalities. Indication of muscle damage as assessed by AST, LDH and CK values was most prominent elevated in protocol 1, 3A, and 3B. CONCLUSIONS: We conclude that intramuscular administration of 12 mg/kg alphaxalone to common marmosets is preferred over other protocols studied. Protocol 2 resulted in at least comparable immobilisation quality with acceptable and less frequent side effects and superior recovery quality. In all protocols, supportive therapy, such as external heat support, remains mandatory. Notably, an unacceptable long recovery period in both ketamine/medetomidine protocols (subsequently reversed with atipamezole) was observed, showing that α-2 adrenoreceptor agonists in the used dose and dosing regime is not the first choice for sedation in common marmosets in a standard research setting.

Reference Intervals and Percentiles for Hematologic and Serum Biochemical Values in Captive Bred Rhesus (Macaca mulatta) and Cynomolgus Macaques (Macaca fascicularis).

Several physiological characteristics and housing conditions are known to affect hematologic and serum biochemical values in macaques. However, the studies that have been conducted either report values calculated based on a small number of animals, were designed specifically to document the effect of a particular condition on the normal range of hematologic and serum biochemical values, or used parametric assumptions to calculate hematologic and serum biochemical reference intervals. We conducted a retrospective longitudinal cohort study to estimate reference intervals for hematologic and serum biochemical values in clinically healthy macaques based on observed percentiles without parametric assumptions. Data were obtained as part of the Biomedical Primate Research Centre (Rijswijk, The Netherlands) health monitoring program between 2018 and 2021. In total, 4009 blood samples from 1475 macaques were analyzed with a maximum of one repeat per year per animal. Data were established by species, gender, age, weight-for-height indices, pregnancy, sedation protocol, and housing conditions. Most of the parameters profoundly affected just some hematologic and serum biochemical values. A significant glucose difference was observed between the ketamine and ketamine-medetomidine sedation protocols. The results emphasize the importance of establishing uniform experimental groups with validated animal husbandry and housing conditions to improve the reproducibility of the experiments.

Long-acting reversible contraception with etonogestrel implants in female macaques (Macaca mulatta and Macaca fascicularis).

Introduction: Contraception is often required for management and population control purposes in group-housed and free-roaming non-human primates. Long-acting reversible contraceptives, including subdermal progestin-releasing implants, are preferred as they eliminate challenges associated with frequent administration. Etonogestrel (ENG)-releasing subdermal implants are reversible and long-acting for a minimum of 3 years, and are commercially available for human use as Implanon® or Nexplanon®. Methods: A retrospective analysis was performed detailing the contraceptive effectiveness and reversibility of subdermal placement of one-fourth or one-third of an ENG implant (68 mg/implant) in 129 female rhesus macaques (Macaca mulatta) and 67 cynomolgus macaques (Macaca fascicularis) at the Biomedical Primate Research Centre (Rijswijk, Netherlands). Furthermore, single cross-sectional ENG serum concentrations were measured for 16 rhesus and 10 cynomolgus macaques, and hemoglobin and blood chemistry pre-ENG and at timepoints >0.5, >1.5, and > 2.5 years post-ENG insertion were evaluated for 24 rhesus macaques. Finally, data were obtained using trans-abdominal ultrasound regarding the influence of ENG on uterine volume and endometrial thickness in 14 rhesus and 11 cynomolgus macaques. Results: As a contraceptive ENG was in 99.80% (CI 93.50-99.99) and 99.95% (CI 99.95-100) effective in rhesus and cynomolgus macaques, respectively. Prolonged ENG durations of implant use in 14 rhesus macaques (range 3.1-5.0 years) and eight cynomolgus macaques (range 3.2-4.0 years) resulted in no unintended pregnancies. A total of 17 female macaques were allowed to breed after ENG removal, and among them, 14 female macaques (82%) had an uneventful delivery. Serum ENG concentrations with a median ENG duration of 1.2 years (range 0.1-6.0 years) and 1.9 years (range 0.6-4.7 years) resulted in median concentrations of 112 pg./mL (range 0-305 pg./mL) and 310 pg./mL (range 183-382 pg./mL) for rhesus and cynomolgus macaques, respectively. ENG had no clinical effect on hemoglobin and blood chemistry parameters nor on the thickness of the endometrial lining or uterus volume. Conclusion: This study indicates that both one-fourth and one-third of the ENG implants are effective, long-acting, reversible, and safe contraceptive to use in macaques.

Potential business model for a European vaccine R&D infrastructure and its estimated socio-economic impact.

Background: Research infrastructures are facilities or resources that have proven fundamental for supporting scientific research and innovation. However, they are also known to be very expensive in their establishment, operation and maintenance. As by far the biggest share of these costs is always borne by public funders, there is a strong interest and indeed a necessity to develop alternative business models for such infrastructures that allow them to function in a more sustainable manner that is less dependent on public financing. Methods: In this article, we describe a feasibility study we have undertaken to develop a potentially sustainable business model for a vaccine research and development (R&D) infrastructure. The model we have developed integrates two different types of business models that would provide the infrastructure with two different types of revenue streams which would facilitate its establishment and would be a measure of risk reduction. For the business model we are proposing, we have undertaken an ex ante impact assessment that estimates the expected impact for a vaccine R&D infrastructure based on the proposed models along three different dimensions: health, society and economy. Results: Our impact assessment demonstrates that such a vaccine R&D infrastructure could achieve a very significant socio-economic impact, and so its establishment is therefore considered worthwhile pursuing. Conclusions: The business model we have developed, the impact assessment and the overall process we have followed might also be of interest to other research infrastructure initiatives in the biomedical field.

Multigenerational Social Housing and Group-Rearing Enhance Female Reproductive Success in Captive Rhesus Macaques (Macaca mulatta).

To optimize costs and reproductive success, rhesus macaques in biomedical primate research facilities are often peer-reared. Older, dependent infants are typically removed from their natal group to enhance female reproduction. The minimal husbandry age-norm of infant removal is ten months. These practices deviate from species-specific behavior and may reduce welfare, suggesting a trade-off between female reproduction and welfare. However, the effect of breeding group type and rearing history on female reproductive success (i.e., birth rate; inter-birth interval (IBI); offspring survival) is unclear. This retrospective study investigated whether group type (i.e., peer groups versus multigenerational groups) and rearing history (i.e., peer- or hand-reared; group-reared with peer- or hand-reared mother; group-reared) affected female reproductive success in captive rhesus macaques. Data on female reproduction between 1996 and 2019 were collected at the Biomedical Primate Research Centre, Rijswijk. Birth rates were higher in multigenerational breeding groups than in peer groups. Moreover, group-reared females had higher offspring survival than peer- or hand-reared females. IBI was not affected by breeding group type or female rearing history. However, females in both peer and multigenerational breeding groups typically conceived earlier after giving birth than the husbandry infant removal age-norm of ten months. Thus, infant removal at an age of ten months does not enhance a female's reproduction. Altogether, female reproduction and non-human primate welfare can simultaneously be optimized through multigenerational breeding groups and group-rearing.

Assessment of Indoor Air Quality for Group-Housed Macaques (Macaca spp.).

Indoor Air Quality (IAQ) is strongly associated with animal health and wellbeing. To identify possible problems of the indoor environment of macaques (Macaca spp.), we assessed the IAQ. The temperature (°C), relative humidity (%) and concentrations of inhalable dust (mg/m3), endotoxins (EU/m3), ammonia (ppm) and fungal aerosols were measured at stationary fixed locations in indoor enclosures of group-housed rhesus (Macaca mulatta) and cynomolgus macaques (Macaca fascicularis). In addition, the personal exposure of caretakers to inhalable dust and endotoxins was measured and evaluated. Furthermore, the air circulation was assessed with non-toxic smoke, and the number of times the macaques sneezed was recorded. The indoor temperature and relative humidity for both species were within comfortable ranges. The geometric mean (GM) ammonia, dust and endotoxin concentrations were 1.84 and 0.58 ppm, 0.07 and 0.07 mg/m3, and 24.8 and 6.44 EU/m3 in the rhesus and cynomolgus macaque units, respectively. The GM dust concentrations were significantly higher during the daytime than during the nighttime. Airborne fungi ranged between 425 and 1877 CFU/m3. Personal measurements on the caretakers showed GM dust and endotoxin concentrations of 4.2 mg/m3 and 439.0 EU/m3, respectively. The number of sneezes and the IAQ parameters were not correlated. The smoke test revealed a suboptimal air flow pattern. Although the dust, endotoxins and ammonia were revealed to be within accepted human threshold limit values (TLV), caretakers were exposed to dust and endotoxin levels exceeding existing occupational reference values.

Overweight Management through Mild Caloric Restriction in Multigenerational Long-Tailed Macaque Breeding Groups.

Caloric restriction (CR) is an effective method to reduce overweight in captive non-human primates (NHPs). CR has been applied to individually- and pair-housed NHPs, but whether applying CR can be effective and safe in group-housed NHPs has not yet been assessed. This study investigates the effect of mild (20%) CR on adult overweight and biochemical parameters, immature growth, veterinary consultations, and reproductive success in multigenerational long-tailed macaque (Macaca fascicularis) breeding groups. Data were derived from anthropometric measurements and blood samples during yearly health checks, complemented with retrospective data on veterinary consultations and reproductive success. Adult body measures decreased after CR, with heavier individuals and females losing more weight compared to leaner individuals and males. CR lowered cholesterol levels in adults but had no overall effect on other biochemical parameters. Yet, biochemical parameters of individuals with high baseline values were reduced more compared to individuals with low baseline values. Immature growth, veterinary consultations and reproductive success were not influenced by CR. Thus, CR targeted the right individuals, i.e., overweight adults, and had no adverse effects on the variables examined in this study. This implies that mild CR can be a valuable overweight management strategy in group-housed NHPs.

Viral biogeography of the mammalian gut and parenchymal organs.

The mammalian virome has been linked to health and disease but our understanding of how it is structured along the longitudinal axis of the mammalian gastrointestinal tract (GIT) and other organs is limited. Here, we report a metagenomic analysis of the prokaryotic and eukaryotic virome occupying luminal and mucosa-associated habitats along the GIT, as well as parenchymal organs (liver, lung and spleen), in two representative mammalian species, the domestic pig and rhesus macaque (six animals per species). Luminal samples from the large intestine of both mammals harboured the highest loads and diversity of bacteriophages (class Caudoviricetes, family Microviridae and others). Mucosal samples contained much lower viral loads but a higher proportion of eukaryotic viruses (families Astroviridae, Caliciviridae, Parvoviridae). Parenchymal organs contained bacteriophages of gut origin, in addition to some eukaryotic viruses. Overall, GIT virome composition was specific to anatomical region and host species. Upper GIT and mucosa-specific viruses were greatly under-represented in distal colon samples (a proxy for faeces). Nonetheless, certain viral and phage species were ubiquitous in all samples from the oral cavity to the distal colon. The dataset and its accompanying methodology may provide an important resource for future work investigating the biogeography of the mammalian gut virome.

Novel application of [18F]DPA714 for visualizing the pulmonary inflammation process of SARS-CoV-2-infection in rhesus monkeys (Macaca mulatta).

RATIONALE: The aim of this study was to investigate the application of [18F]DPA714 to visualize the inflammation process in the lungs of SARS-CoV-2-infected rhesus monkeys, focusing on the presence of pulmonary lesions, activation of mediastinal lymph nodes and surrounded lung tissue. METHODS: Four experimentally SARS-CoV-2 infected rhesus monkeys were followed for seven weeks post infection (pi) with a weekly PET-CT using [18F]DPA714. Two PET images, 10 min each, of a single field-of-view covering the chest area, were obtained 10 and 30 min after injection. To determine the infection process swabs, blood and bronchoalveolar lavages (BALs) were obtained. RESULTS: All animals were positive for SARS-CoV-2 in both the swabs and BALs on multiple timepoints pi. The initial development of pulmonary lesions was already detected at the first scan, performed 2-days pi. PET revealed an increased tracer uptake in the pulmonary lesions and mediastinal lymph nodes of all animals from the first scan obtained after infection and onwards. However, also an increased uptake was detected in the lung tissue surrounding the lesions, which persisted until day 30 and then subsided by day 37-44 pi. In parallel, a similar pattern of increased expression of activation markers was observed on dendritic cells in blood. PRINCIPAL CONCLUSIONS: This study illustrates that [18F]DPA714 is a valuable radiotracer to visualize SARS-CoV-2-associated pulmonary inflammation, which coincided with activation of dendritic cells in blood. [18F]DPA714 thus has the potential to be of added value as diagnostic tracer for other viral respiratory infections.

Brain Inflammation and Intracellular α-Synuclein Aggregates in Macaques after SARS-CoV-2 Infection.

SARS-CoV-2 causes acute respiratory disease, but many patients also experience neurological complications. Neuropathological changes with pronounced neuroinflammation have been described in individuals after lethal COVID-19, as well as in the CSF of hospitalized patients with neurological complications. To assess whether neuropathological changes can occur after a SARS-CoV-2 infection, leading to mild-to-moderate disease, we investigated the brains of four rhesus and four cynomolgus macaques after pulmonary disease and without overt clinical symptoms. Postmortem analysis demonstrated the infiltration of T-cells and activated microglia in the parenchyma of all infected animals, even in the absence of viral antigen or RNA. Moreover, intracellular α-synuclein aggregates were found in the brains of both macaque species. The heterogeneity of these manifestations in the brains indicates the virus' neuropathological potential and should be considered a warning for long-term health risks, following SARS-CoV-2 infection.

Medical imaging of pulmonary disease in SARS-CoV-2-exposed non-human primates.

Chest X-ray (CXR), computed tomography (CT), and positron emission tomography-computed tomography (PET-CT) are noninvasive imaging techniques widely used in human and veterinary pulmonary research and medicine. These techniques have recently been applied in studies of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-exposed non-human primates (NHPs) to complement virological assessments with meaningful translational readouts of lung disease. Our review of the literature indicates that medical imaging of SARS-CoV-2-exposed NHPs enables high-resolution qualitative and quantitative characterization of disease otherwise clinically invisible and potentially provides user-independent and unbiased evaluation of medical countermeasures (MCMs). However, we also found high variability in image acquisition and analysis protocols among studies. These findings uncover an urgent need to improve standardization and ensure direct comparability across studies.