Preclinical in vivo assessment of the activity of AZD7442 anti-SARS-CoV-2 monoclonal antibodies against Omicron sublineages.

Therapeutic monoclonal antibodies have been successful in protecting vulnerable populations against SARS-CoV-2. However, their effectiveness has been hampered by the emergence of new variants. To adapt the therapeutic landscape, health authorities have based their recommendations mostly on in vitro neutralization tests. However, these do not provide a reliable understanding of the changes in the dose-effect relationship and how they may translate into clinical efficacy. Taking the example of EvusheldTM (AZD7442), we aimed to investigate how in vivo data can provide critical quantitative results and project clinical effectiveness. We used the Golden Syrian hamster model to estimate 90 % effective concentrations (EC90) of AZD7442 in vivo against SARS-CoV-2 Omicron BA.1, BA.2 and BA.5 variants. While our in vivo results confirmed the partial loss of AZD7442 activity for BA.1 and BA.2, they showed a much greater loss of efficacy against BA.5 than that obtained in vitro. We analyzed in vivo EC90s in perspective with antibody levels measured in a cohort of immunocompromised patients who received 300 mg of AZD7442. We found that a substantial proportion of patients had serum levels of anti-SARS-CoV-2 spike protein IgG above the estimated in vivo EC90 for BA.1 and BA.2 (21 % and 92 % after 1 month, respectively), but not for BA.5. These findings suggest that AZD7442 is likely to retain clinical efficacy against BA.2 and BA.1, but not against BA.5. Overall, the present study illustrates the importance of complementing in vitro investigations by preclinical studies in animal models to help predict the efficacy of monoclonal antibodies in humans.

Broad-spectrum dengue virus detection using the commercial RealStar dengue RT-PCR kit 3.0 (Altona) and an in-house combined real-time RT-PCR assay.

In endemic areas, the genetic diversity among co-circulating dengue virus (DENV) strains is considerable and new, highly divergent strains are identified on a regular basis. It is thus critical to ensure that molecular diagnostic tools effectively detect virus genomes even in case of important genetic variation. Here, we tested both the pan-DENV detection capacity and the limit of detection of two real-time RT-PCR assays: (i) the commercial RealStar Altona 3.0 system and (ii) a laboratory developed test (LDT) combining two RT-PCR systems in a single reaction tube (DenAllDUO). We used a panel of DENV strains representative of the genetic diversity within DENV species, combined with three in vitro transcribed RNAs as surrogates for unavailable strains corresponding to recently discovered strains with substantial genetic divergence: DENV serotype 1 (DENV-1) Brun2014, DENV-2 QML22 and DENV-4 DKE121. Both systems (i) targeted the genome 3' untranslated region, (ii) displayed a broad detection spectrum, encompassing most of DENV species diversity, and (iii) detected the three aforementioned divergent strains. DenAllDUO detected all the strains tested, whereas the RealStar system failed to detect strains from DENV-4 genotype III. Altogether, our findings support the value of these two RT-PCR systems as part of the Dengue diagnostic arsenal.

Further preclinical characterization of molnupiravir against SARS-CoV-2: Antiviral activity determinants and viral genome alteration patterns.

The SARS-CoV-2 pandemic has highlighted the need for broad-spectrum antiviral drugs to respond promptly to viral emergence. We conducted a preclinical study of molnupiravir (MOV) against SARS-CoV-2 to fully characterise its antiviral properties and mode of action. The antiviral activity of different concentrations of MOV was evaluated ex vivo on human airway epithelium (HAE) and in vivo in a hamster model at three escalating doses (150, 300 and 400 mg/kg/day) according to three different regimens (preventive, pre-emptive and curative). We assessed viral loads and infectious titres at the apical pole of HAE and in hamster lungs, and MOV trough concentration in plasma and lungs. To explore the mode of action of the MOV, the entire genomes of the collected viruses were deep-sequenced. MOV effectively reduced viral titres in HAE and in the lungs of treated animals. Early treatment after infection was a key factor in efficacy, probably associated with high lung concentrations of MOV, suggesting good accumulation in the lung. MOV induced genomic alteration in viral genomes with an increase in the number of minority variants, and predominant G to A transitions. The observed reduction in viral replication and its mechanism of action leading to lethal mutagenesis, supported by clinical trials showing antiviral action in humans, provide a convincing basis for further research as an additional means in the fight against COVID-19 and other RNA viruses.

Bacterial Agents Detected in 418 Ticks Removed from Humans during 2014-2021, France.

Monitoring of tickborne diseases is critical for prevention and management. We analyzed 418 ticks removed from 359 patients during 2014-2021 in Marseille, France, for identification and bacteria detection. Using morphology, molecular methods, or matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, we identified 197 (47%) Ixodes, 136 (33%) Dermacentor, 67 (16%) Rhipicephalus, 8 (2%) Hyalomma, 6 (1%) Amblyomma, 2 (0.5%) Argas, and 2 (0.5%) Haemaphysalis tick species. We also detected bacterial DNA in 241 (58%) ticks. The most frequent bacterial pathogens were Rickettsia raoultii (17%) and R. slovaca (13%) in Dermacentor ticks, Borrelia spp. (9%) in Ixodes ticks, and R. massiliae (16%) in Rhipicephalus ticks. Among patients who were bitten, 107 had symptoms, and tickborne diseases were diagnosed in 26, including scalp eschar and neck lymphadenopathy after tick bite and Lyme borrelioses. Rapid tick and bacteria identification using a combination of methods can substantially contribute to clinical diagnosis, treatment, and surveillance of tickborne diseases.

Orthopoxvirus Seroprevalence and Infection Susceptibility in France, Bolivia, Laos, and Mali.

To determine a demographic overview of orthopoxvirus seroprevalence, we tested blood samples collected during 2003-2019 from France (n = 4,876), Bolivia (n = 601), Laos (n = 657), and Mali (n = 255) for neutralizing antibodies against vaccinia virus. In addition, we tested 4,448 of the 4,876 samples from France for neutralizing antibodies against cowpox virus. We confirmed extensive cross-immunity between the 2 viruses. Seroprevalence of antibodies was <1% in Bolivia, <5% in Laos, and 17.25% in Mali. In France, we found low prevalence of neutralizing antibodies in persons who were unvaccinated and vaccinated for smallpox, suggesting immunosenescence occurred in vaccinated persons, and smallpox vaccination compliance declined before the end of compulsory vaccination. Our results suggest that populations in Europe, Africa, Asia, and South America are susceptible to orthopoxvirus infections, which might have precipitated the emergence of orthopoxvirus infections such as the 2022 spread of monkeypox in Europe.

Detection of Cetacean Poxvirus in Peruvian Common Bottlenose Dolphins (Tursiops truncatus) Using a Pan-Poxvirus PCR.

Cetacean poxviruses (CePVs) cause 'tattoo' skin lesions in small and large cetaceans worldwide. Although the disease has been known for decades, genomic data for these poxviruses are very limited, with the exception of CePV-Tursiops aduncus, which was completely sequenced in 2020. Using a newly developed pan-pox real-time PCR system targeting a conserved nucleotide sequence located within the Monkeypox virus D6R gene, we rapidly detected the CePV genome in typical skin lesions collected from two Peruvian common bottlenose dolphins (Tursiops truncatus) by-caught off Peru in 1993. Phylogenetic analyses based on the sequencing of the DNA polymerase and DNA topoisomerase genes showed that the two viruses are very closely related to each other, although the dolphins they infected pertained to different ecotypes. The poxviruses described in this study belong to CePV-1, a heterogeneous clade that infects many species of dolphins (Delphinidae) and porpoises (Phocoenidae). Among this clade, the T. truncatus CePVs from Peru were more related to the viruses infecting Delphinidae than to those detected in Phocoenidae. This is the first time that CePVs were identified in free-ranging odontocetes from the Eastern Pacific, surprisingly in 30-year-old samples. These data further suggest a close and long-standing pathogen-host co-evolution, resulting in different lineages of CePVs.

Need for a Standardized Translational Drug Development Platform: Lessons Learned from the Repurposing of Drugs for COVID-19.

In the absence of drugs to treat or prevent COVID-19, drug repurposing can be a valuable strategy. Despite a substantial number of clinical trials, drug repurposing did not deliver on its promise. While success was observed with some repurposed drugs (e.g., remdesivir, dexamethasone, tocilizumab, baricitinib), others failed to show clinical efficacy. One reason is the lack of clear translational processes based on adequate preclinical profiling before clinical evaluation. Combined with limitations of existing in vitro and in vivo models, there is a need for a systematic approach to urgent antiviral drug development in the context of a global pandemic. We implemented a methodology to test repurposed and experimental drugs to generate robust preclinical evidence for further clinical development. This translational drug development platform comprises in vitro, ex vivo, and in vivo models of SARS-CoV-2, along with pharmacokinetic modeling and simulation approaches to evaluate exposure levels in plasma and target organs. Here, we provide examples of identified repurposed antiviral drugs tested within our multidisciplinary collaboration to highlight lessons learned in urgent antiviral drug development during the COVID-19 pandemic. Our data confirm the importance of assessing in vitro and in vivo potency in multiple assays to boost the translatability of pre-clinical data. The value of pharmacokinetic modeling and simulations for compound prioritization is also discussed. We advocate the need for a standardized translational drug development platform for mild-to-moderate COVID-19 to generate preclinical evidence in support of clinical trials. We propose clear prerequisites for progression of drug candidates for repurposing into clinical trials. Further research is needed to gain a deeper understanding of the scope and limitations of the presented translational drug development platform.

Atypical Borrelia garinii infection in an immunocompromised patient mimicking high-grade lymphoma.

We report an atypical Borrelia garinii infection in a patient who was immunocompromised. It was first suspected as a transformation of follicular lymphoma into high-grade lymphoma. Spirochetes were directly observed on a peripheral blood smear and the diagnosis was confirmed using molecular methods. The clinical presentation and the diagnosis are unique and contrast with the cases described in the literature in patients who are immunocompromised.

The SARS-CoV-2 Alpha variant exhibits comparable fitness to the D614G strain in a Syrian hamster model.

Late 2020, SARS-CoV-2 Alpha variant emerged in United Kingdom and gradually replaced G614 strains initially involved in the global spread of the pandemic. In this study, we use a Syrian hamster model to compare a clinical strain of Alpha variant with an ancestral G614 strain. The Alpha variant succeed to infect animals and to induce a pathology that mimics COVID-19. However, both strains replicate to almost the same level and induced a comparable disease and immune response. A slight fitness advantage is noted for the G614 strain during competition and transmission experiments. These data do not corroborate the epidemiological situation observed during the first half of 2021 in humans nor reports that showed a more rapid replication of Alpha variant in human reconstituted bronchial epithelium. This study highlights the need to combine data from different laboratories using various animal models to decipher the biological properties of newly emerging SARS-CoV-2 variants.

Serious and Atypical Presentations of Bartonella henselae Infection in Kidney Transplant Recipients.

This article describes 5 cases of bartonellosis with fever and atypical clinical presentations in kidney transplant recipients: thrombotic microangiopathies, recurrent hemophagocytosis, and immune reconstitution syndrome after treatment. The diagnosis, the pathological lesions, and treatments are described. Bartonellosis must be researched in solid organ transplant recipients with fever of undetermined origin.

Molecular detection of parapoxvirus in Ixodidae ticks collected from cattle in Corsica, France.

BACKGROUND: Several viruses belonging to the family Poxviridae can cause infections in humans and animals. In Corsica, livestock farming (sheep, goats, pigs, and cattle) is mainly mixed, leading to important interactions between livestock, wildlife, and human populations. This could facilitate the circulation of zoonotic diseases, and makes Corsica a good example for studies of tick-borne diseases. OBJECTIVES: To gain understanding on the circulation of poxviruses in Corsica, we investigated their presence in tick species collected from cattle, sheep, horses, and wild boar, and characterized them through molecular techniques. METHODS: Ticks were tested using specific primers targeting conserved regions of sequences corresponding to two genera: parapoxvirus and orthopoxvirus. RESULTS: A total of 3555 ticks were collected from 1549 different animals (687 cattle, 538 horses, 106 sheep, and 218 wild boars). They were tested for the presence of parapoxvirus DNA on one hand and orthopoxvirus DNA on the other hand using Pangeneric real-time TaqMan assays. Orthopoxvirus DNA was detected in none of the 3555 ticks. Parapoxvirus DNA was detected in 6.6% (36/544) of ticks collected from 23 cows from 20 farms. The remaining 3011 ticks collected from horses, wild boars, and sheep were negative. The infection rate in cow ticks was 8.0% (12/148) in 2018 and 6.0% (24/396) in 2019 (p = 0.57). Parapoxvirus DNA was detected in 8.5% (5/59) of Hyalomma scupense pools, 8.2% (15/183) of Hyalomma marginatum pools, and 6.7% (16/240) of Rhipicephalus bursa pools (p = 0.73). We successfully amplified and sequenced 19.4% (7/36) of the positive samples which all corresponded to pseudocowpox virus. CONCLUSIONS: Obviously, further studies are needed to investigate the zoonotic potential of pseudocowpox virus and its importance for animals and public health.

Similar patterns of [18F]-FDG brain PET hypometabolism in paediatric and adult patients with long COVID: a paediatric case series.

PURPOSE: Several weeks after COVID-19 infection, some children report the persistence or recurrence of functional complaints. This clinical presentation has been referred as "long COVID" in the adult population, and an [18F]-FDG brain PET hypometabolic pattern has recently been suggested as a biomarker. Herein, we present a retrospective analysis of 7 paediatric patients with suspected long COVID who were explored by [18F]-FDG brain PET exam. Metabolic brain findings were confronted to those obtained in adult patients with long COVID, in comparison to their respective age-matched control groups. METHODS: Review of clinical examination and whole-brain voxel-based analysis of [18F]-FDG PET metabolism of the 7 children in comparison to 21 paediatric controls, 35 adult patients with long COVID and 44 healthy adult subjects. RESULTS: Despite lower initial severity at the acute stage of the infection, paediatric patients demonstrated on average 5 months later a similar brain hypometabolic pattern as that found in adult long COVID patients, involving bilateral medial temporal lobes, brainstem and cerebellum (p-voxel < 0.001, p-cluster < 0.05 FWE-corrected), and also the right olfactory gyrus after small volume correction (p-voxel = 0.010 FWE-corrected), with partial PET recovery in two children at follow-up. CONCLUSION: These results provide arguments in favour of possible long COVID in children, with a similar functional brain involvement to those found in adults, regardless of age and initial severity.

Pneumococcal pneumonia among shipyard workers: Inside the features of disease onset.

BACKGROUND: Several outbreaks of pneumococcal pneumonia among shipyard workers have been described. In this study, following a previous report of grouped cases, we aimed to elucidate the features of disease onset. METHODS: We compared the population characteristics of shipyard workers with a confirmed diagnosis of pneumococcal pneumonia (N = 38) to those of workers without pneumonia (N = 53). We compared nine S. pneumoniae strains isolated from patients with pneumonia by capsular serotyping, multi-locus sequence typing, and whole genome sequencing. RESULTS: Shipyard workers with Streptococcus pneumoniae pneumonia were more frequently from Italy (P = 0.016), had at least one underlying condition (P = 0.024), lived on-board the ship (P = 0.009). None of these factors was independent by multivariate analysis. While capsular serotyping enabled us to identify four different serotypes: 4 (n = 5), 8 (n = 2), 9 N (n = 1), and 3 (n = 1), by sequence typing, we distinguished five sequence types (STs): ST801 (n = 4), ST205 (n = 2), ST1220 (n = 1), ST1280 (n = 1), and ST66 (n = 1). Whole genome sequencing confirmed the results obtained by MLST. Genomes of isolates of the same sequence type were similar with ≤80 single-nucleotide polymorphisms. CONCLUSIONS: We confirmed that the onset of pneumococcal infection among shipyard workers was attributable to both a person-to-person spread of single strains of S. pneumoniae and a shift of different strains from commensal to pathogen under favourable conditions (professional exposure, viral infections). Control measures should therefore be implemented by taking into account these features.

Measurement of the IgG Avidity Index in the Diagnosis of Clinical Toxocariasis Patients.

Toxocara spp. are parasitic nematodes responsible for human toxocariasis, a common zoonotic helminth infection. The five main features of human toxocariasis are the classical ocular toxocariasis and visceral larva migrans syndrome, followed by covert toxocariasis, common toxocariasis and neurotoxocariasis. The diagnosis of toxocariasis is feasible by considering clinical symptoms, anamnestic history and serology laboratory results; however, serological criteria cannot be used to distinguish active Toxocara infection from past exposure, which is an area of much discussion in clinical practice. In this context, we developed avidity tests (ELISA and immunoblotting) and evaluated their clinical usefulness in distinguishing past from active toxocariasis. Our study involved 46 patients divided into two groups: "active toxocariasis" (n = 14) and "chronic toxocariasis" (n = 32). According to the avidity indices obtained for both the chronic and active toxocariasis groups, we proposed two thresholds: first, an AI lower than 32% supports an active infection; secondly, a threshold above 42% can exclude an active infection. In order to use this assay in routine clinical practice, however, is still requires standardisation with regards to the method and threshold values, which can be established through studies involving larger populations.

Investigation of Bufavirus and Parvovirus 4 in Patients with Gastro-Enteritis from the South-East of France.

Bufavirus (BuV) and human parvovirus 4 (PARV4) belong to the Parvoviridae family. We assessed BuV and PARV4 DNA presence by real-time PCR analysis in stool, blood and respiratory samples collected in patients from Marseille and Nice, two large cities in the South-East of France. Bu-V DNA was detected in diarrheic stool samples from 92 patients (3.6% of 2583 patients), particularly men and adults, and patients from the nephrology and the infectious disease departments. Among the patients with a BuV-positive stool sample and for whom at least one blood sample was available (n = 30 patients), BuV DNA was detected also in 3 blood samples. In contrast, BuV DNA was not detected in any of the respiratory samples from 23 patients with BuV-positive stool. BuV detection rate was comparable in stool samples from patients with and without diarrhea. We did not detect PARV4 DNA in any of the stool specimens (n = 2583 patients). Our results suggest that PARV4 fecal-oral transmission is rare or non-existent in the South-East of France while BuV circulates with a relatively high rate in this area.

Evaluation of the Ridaquick Rotavirus/Adenovirus Immuno-Chromatographic Assay in Real-Life Situation.

Immunochromatographic tests (ICT) are diagnostics tools providing rapid results without the need for specialized equipment. Our aim was to evaluate retrospectively the rotavirus and adenovirus ICT routinely used in the virology laboratory serving the University Hospital of Marseille, France. From January 2017 to March 2020, 715 stool specimens from patients were screened using the Ridaquick Rotavirus/Adenovirus Combi ICT (RR/AC ICT) and a commercially available multiplex PCR detection kit. Rotavirus was detected in 9.2% of specimens by PCR and 7.7% of specimens by RR/AC ICT while adenovirus was detected in 8.5% of specimens by PCR and 2.4% of specimens by RR/AC ICT. The RR/AC ICT parameters for rotavirus were 75.8% sensitivity, 99.2% specificity, 90.9% positive predictive value (PPV) and 97.6% negative predictive value (NPV). The RR/AC ICT parameters for adenovirus were 6.6% sensitivity, 98.0% specificity, 23.5% PPV and 91.8% NPV. While the ICT test may be suitable for rotavirus detection, a PCR-based assay is better adapted for adenovirus detection in stools.

Detection of Pneumocystis jirovecii in Hospitalized Children Less Than 3 Years of Age.

Few data are available in the literature regarding Pneumocystis jirovecii infection in children under 3 years old. This retrospective cohort study aimed to describe medically relevant information among them. All children under 3 years old treated in the same medical units from April 2014 to August 2020 and in whom a P. jirovecii evaluation was undertaken were enrolled in the study. A positive case was defined as a child presenting at least one positive PCR for P. jirovecii in a respiratory sample. Medically relevant information such as demographical characteristics, clinical presentation, microbiological co-infections, and treatments were collected. The objectives were to describe the characteristics of these children with P. jirovecii colonization/infection to determine the key underlying diseases and risk factors, and to identify viral respiratory pathogens associated. The PCR was positive for P. jirovecii in 32 children. Cardiopulmonary pathologies (21.9%) were the most common underlying disease in them, followed by severe combined immunodeficiency (SCID) (18.8%), hyaline membrane disease (15.6%), asthma (9.4%) and acute leukaemia (6.3%). All SCID children were diagnosed with pneumocystis pneumonia. Co-infection with Pj/Rhinovirus (34.4%) was not significant. Overall mortality was 18.8%. Paediatric pneumocystis is not restricted to patients with HIV or SCID and should be considered in pneumonia in children under 3 years old.