Detection and Molecular Characterization of Canine Alphacoronavirus in Free-Roaming Dogs, Bangladesh

Canine coronavirus (CCoV) is widespread among the dog population and causes gastrointestinal disorders, and even fatal cases. As the zoonotic transmission of viruses from animals to humans has become a worldwide concern nowadays, it is necessary to screen free-roaming dogs for their common pathogens due to their frequent interaction with humans. We conducted a cross-sectional study to detect and characterize the known and novel Corona, Filo, Flavi, and Paramyxoviruses in free-roaming dogs in Bangladesh. Between 2009–10 and 2016–17, we collected swab samples from 69 dogs from four districts of Bangladesh, tested using RT-PCR and sequenced. None of the samples were positive for Filo, Flavi, and Paramyxoviruses. Only three samples (4.3%; 95%CI: 0.9–12.2) tested positive for Canine Coronavirus (CCoV). The CCoV strains identified were branched with strains of genotype CCoV-II with distinct distances. They are closely related to CCoVs from the UK, China, and other CoVs isolated from different species, which suggests genetic recombination and interspecies transmission of CCoVs. These findings indicate that CCoV is circulating in dogs of Bangladesh. Hence, we recommend future studies on epidemiology and genetic characterization with full-genome sequencing of emerging coronaviruses in companion animals in Bangladesh.

Molecular Characterization of Circulating DENV-2 During Outbreak in Northern Senegal, Rosso 2018

Globally 390 millions of people are at risk of dengue infection; over the past 50 years the virus incidence increased thirty-fold. In Senegal, an unprecedented occurrence of outbreaks and sporadic cases was noticed since 2017. In October 2018 an outbreak of DENV-2 was reported in Rosso area in the north of Senegal at the border with Mauritania. Out of the 187 blood specimen samples collected, 27 were positives by qRT-PCR and 8 were serologically positive for DENV IgM. Serotyping using qRT-PCR reveals that isolates were positive for DENV-2. A subset of DENV-2 positives samples was selected and subjected to full genome sequencing followed by phylogenetic analysis. Analysis of 06 nearly completed genome sequences (n= 6) revealed that isolates belong to the cosmopolitan genotype and are closely related to the Mauritanian strains detected between 2017 and 2018 and those detected in many West African countries such as Burkina Faso or Cote d’Ivoire. Our results suggest a transboundary circulation of the DENV-2 cosmopolitan genotype between Senegal and Mauritania and call for a need of coordinated surveillance of arboviruses between these two countries. Interestingly, high level of homology between West African isolates highlights endemicity and call for a set-up of sub-regional viral genomic surveillance which will lead to a better understanding of viral dynamic, transmission and spread across Africa.

Case Series: Convalescent Plasma Therapy for Patients with COVID-19 and Primary Antibody Deficiency

Patients with primary antibody deficiency are at risk for severe and in many cases for prolonged COVID-19. Convalescent plasma treatment of immunocompromised individuals could be an option especially in countries with limited access to monoclonal antibody therapies. While studies in immunocompetent COVID19 patients have demonstrated only a limited benefit, evidence for the safety, timing, and effectiveness of this treatment in antibody-deficient patients is lacking. Here, we describe 16 cases with primary antibody deficiency treated with convalescent plasma in four medical centers. In our cohort, treatment was associated with a reduction in viral load and improvement of clinical symptoms, even when applied over a week after onset of infection. There were no relevant side effects besides a short-term fever reaction in one patient. Longitudinal full-genome sequencing revealed the emergence of mutations in the viral genome, potentially conferring an antibody escape in one patient with persistent viral RNA shedding upon plasma treatment. However, he resolved the infection after a second course of plasma treatment. Thus, our data suggest a therapeutic benefit of convalescent plasma treatment in patients with primary antibody deficiency even months after infection. While it appears to be safe, PCR follow-up for SARS-CoV-2 is advisable and early re-treatment might be considered in patients with persistent viral shedding.

Emergence of B.1.524(G) SARS-CoV-2 in Malaysia during the third COVID-19 epidemic wave

The COVID-19 pandemic first emerged in Malaysia in Jan 2020. As of 12th Sept 2021, 1,979,698 COVID-19 cases that occurred over three major epidemic waves were confirmed. The virus contributing to the three epidemic waves has not been well-studied. We sequenced the genome of 22 SARS-CoV-2 strains detected in Malaysia during the second and the ongoing third wave of the COVID-19 epidemic. Detailed phylogenetic and genetic variation analyses of the SARS-CoV-2 isolate genomes were performed using these newly determined sequences and all other available sequences. Results from the analyses suggested multiple independent introductions of SARS-CoV-2 into Malaysia. A new B.1.524(G) lineage with S-D614G mutation was detected in Sabah, East Malaysia and Selangor, Peninsular Malaysia on 7th October 2020 and 14th October 2020, respectively. This new B.1.524(G) group was not the direct descendant of any of the previously detected lineages. The new B.1.524(G) carried a set of genetic variations, including A701V (position variant frequency = 0.0007) in Spike protein and a novel G114T mutation at the 5’UTR. The biological importance of the specific mutations remained unknown. The sequential appearance of the mutations, however, suggests that the spread of the new B.1.524(G) lineages likely begun in Sabah and then spread to Selangor. The findings presented here support the importance of SARS-CoV-2 full genome sequencing as a tool to establish an epidemiological link between cases or clusters of COVID-19 worldwide.

A novel and highly divergent Canine Distemper Virus lineage causing distemper in ferrets in Australia

Canine distemper virus (CDV) is a highly contagious systemic viral disease of dogs, that regularly spills-over into other animal species. Despite widespread vaccination, CDV remains endemic in many parts of the world. In this study we report an outbreak of distemper in ferrets in two independent research facilities in Australia. We found that disease severity varied, although most animals had mild to moderate disease signs. Histopathology results of animals with severe disease presented the typical profile of distemper pathology with multi-system virus replication. Through the development of a discriminatory PCR paired with full genome sequencing we revealed that the outbreak at both facilities was caused by a single, novel lineage of CDV. This lineage was highly divergent across the H gene, F signal peptide and full genome and had less than 93% similarity across the H gene to other described lineages, including the vaccine strain. Molecular analysis indicates that this strain belongs to a distinct lineage that diverged from other clades approximately 140 to 400 years ago, and appears to be unique to Australia. Given the differences in key viral proteins of this novel CDV strain, a review of the efficacy of the CDV vaccines currently in use in Australia is warranted to ensure maximum protection of dogs and other vulnerable species. In addition, enhanced surveillance to determine the prevalence of CDV in ferrets, dogs and other at-risk species in Australia would be useful to better understand the diversity of CDV in Australia.

Outbreak of SARS-CoV-2 B.1.617.2 (Delta) variant in a Nursing Home 28 weeks after two doses of mRNA anti-Covid-19 vaccines: evidence of a waning immunity

Objectives: Description of a SARS-CoV-2 B.1.617.2 (Delta) variant outbreak among residents (N = 69) and Health Workers (HWs: N = 69) of a small Nursing Home in Northern-East Italy, with full vaccination coverage of 91 and 82 %, respectively. Evaluation of the Anti-Spike IgG titers 28 weeks after the mRNA vaccine boosts against SARS-COV-2 infection and severe Covid-19. Materials and methods: A timely collection of sera within 48 h from the index case; anti-Spike IgG determination (expressed as Binding Antibody Units - BAU/mL) through a commercial quantitative assay; SARS-CoV-2 diagnostics via RT-PCR, and full-genome sequencing for lineage characterization. Residents were grouped according to anti-Spike IgG titers (≤50, 51-1000, and >1000 BAU/mL) and resulting protection against the infection and the severe disease was measured. Results: 0/20 HWs and 14/59 (24 %) residents fully vaccinated and without a previous SARS-CoV-2 infection showed anti-Spike IgG ≤50 BAU/mL (1-sided Fisher exact p=0.011). Among these residents, a level of anti-Spike IgG ≤50 BAU/mL resulted in a higher risk of SARS-CoV-2 infection (RR=1.55, CI 95% 1.17-2.05) and severe Covid-19 disease (RR=5.33, CI95% 1.83-15.57). Conclusion Low levels of SARS-CoV-2 neutralizing anti-Spike IgG in serum 28 weeks after the administration of the second dose parallels the waning of vaccine protection.

Genomic analysis of distinct bleaching tolerances among cryptic coral species

Reef-building coral species are experiencing an unprecedented decline owing to increasing frequency and intensity of marine heatwaves and associated bleaching-induced mortality. Closely related species from the Acropora hyacinthus species complex differ in heat tolerance and in their association with heat-tolerant symbionts. We used low-coverage full genome sequencing of 114 colonies monitored across the 2015 bleaching event in American Samoa to determine the genetic differences among four cryptic species (termed HA, HC, HD and HE) that have diverged in these species traits. Cryptic species differed strongly at thousands of single nucleotide polymorphisms across the genome which are enriched for amino acid changes in the bleaching-resistant species HE. In addition, HE also showed two particularly divergent regions with strong signals of differentiation. One approximately 220 kb locus, HES1, contained the majority of fixed differences in HE. A second locus, HES2, was fixed in HE but polymorphic in the other cryptic species. Surprisingly, non-HE individuals with HE-like haplotypes at HES2 were more likely to bleach. At both loci, HE showed particular sequence similarity to a congener, Acropora millepora . Overall, resilience to bleaching during the third global bleaching event was strongly structured by host cryptic species, buoyed by differences in symbiont associations between these species.

Whole genome sequencing of two human rhinovirus A types (A101 and A15) detected in Kenya, 2016-2018

Background: Virus genome sequencing is increasingly utilized in epidemiological surveillance. Genomic data allows comprehensive evaluation of underlying viral diversity and epidemiology to inform control. For human rhinovirus (HRV), genomic amplification and sequencing is challenging due to numerous types, high genetic diversity and inadequate reference sequences. Methods: We developed a tiled amplicon type-specific protocol for genome amplification and sequencing on the Illumina MiSeq platform of two HRV types, A15 and A101. We then assessed added value in analyzing whole genomes relative to the VP4/2 region only in the investigation of HRV molecular epidemiology within the community in Kilifi, coastal Kenya. Results: We processed 73 nasopharyngeal swabs collected between 2016-2018, and 48 yielded at least 70% HRV genome coverage. These included all A101 samples (n=10) and 38 (60.3%) A15 samples. Phylogenetic analysis revealed that the Kilifi A101 sequences interspersed with global A101 genomes available in GenBank collected between 1999-2016. On the other hand, our A15 sequences formed a monophyletic group separate from the global genomes collected in 2008 and 2019. An improved phylogenetic resolution was observed with the genome phylogenies compared to the VP4/2 phylogenies. Conclusions: We present a type-specific full genome sequencing approach for obtaining HRV genomic data and characterizing infections.

Spread of SARS-CoV-2 Delta variant infections bearing the S:E484Q and S:T95I mutations in July and August 2021 in France

Analysing 92,598 variant screening tests performed on SARS-CoV-2 positive samples collected in France between 1 July and 31 August 2021 shows an increase of Kappa-like infections. Full genome sequencing reveals that these correspond to Delta variants bearing the S:E484Q mutation. Most of these sequences belong to a phylogenetic cluster and also bear the S:T95I mutation. Further monitoring is needed to determine if this trend is driven by undocumented superspreading events or an early signal of future viral evolutionary dynamics

Molecular Confirmation of Ranavirus Infection in Amphibians From Chad, Africa

Ranaviruses are DNA viruses (Family Iridoviridae; Subfamily Alphairidovirinae) and ranaviral disease is considered an emerging infectious disease of ectothermic vertebrates. Ranavirus infection can have varying pathological effects on infected amphibians, reptiles, and fish, most notably causing significant mortality events and population declines. Despite having a broad global range with reports from six continents, only a single incidental finding in Xenopus longipes from mainland Africa (Cameroon) is known and lacks molecular confirmation. Thus, there is a considerable knowledge gap concerning ranaviruses in Africa. We opportunistically obtained tissue samples from 160 amphibians representing five genera (Hoplobatrachus, Hylarana, Ptychadena, Pyxicephalus, and Xenopus) and two turtles (Pelomedusa sp.) from Chad, Africa. Samples were tested for ranavirus infection using a conventional PCR assay targeting the major capsid protein (MCP). A total of 25/160 (16%) frogs tested positive including 15/87 (17%) Hoplobatrachus occipitalis, 10/58 (17%) Ptychadena spp., 0/3 Pyxicephalus spp., 0/9 Xenopus spp., and 0/3 Hylarana spp. One of two turtles tested positive. Partial MCP gene sequences indicated all samples were >98% similar to several frog virus 3 (FV3)-like sequences. Additional gene targets (DNA polymerase [DNApol], ribonucleotide reductase alpha [RNR- α], ribonucleotide reductase beta subunit [RNR- β]) were sequenced to provide further detailed classification of the virus. Sequences of individual gene targets indicate that the ranavirus detected in frogs in Chad is most similar to tiger frog virus (TFV), a FV3-like virus previously isolated from diseased amphibians cultured in China and Thailand. Full genome sequencing of one sample indicates that the Chad frog virus (CFV) is a well-supported sister group to the TFVs previously determined from Asia. This work represents the first molecular confirmation of ranaviruses from Africa and is a first step in comparing ranavirus phylogeography on a local and global scale.