HIV cure: The daunting scale of the problem.

Cure strategies are confounded by basic reservoir biology.

AKT signaling modulates latent viral reservoir viability in HIV-1-infected blood-brain barrier pericytes.

Despite antiretroviral therapy (ART), chronic forms of HIV-associated neurocognitive disorders (HAND) affect an estimated 50% of individuals living with HIV, greatly impacting their quality of life. The prevailing theory of HAND progression posits that chronic inflammation arising from the activation of latent viral reservoirs leads to progressive damage in the central nervous system (CNS). Recent evidence indicates that blood-brain barrier (BBB) pericytes are capable of active HIV-1 infection; however, their latent infection has not been defined. Given their location and function, BBB pericytes are poised to be a key viral reservoir in the development of HAND. We present the first transcriptional analysis of uninfected, active, and latent human BBB pericytes, revealing distinct transcriptional phenotypes. In addition, we demonstrate that latent infection of BBB pericytes relies on AKT signaling for reservoir survival. These findings provide insight into the state of reservoir maintenance in the CNS during HIV-1 infection and provide novel targets for reservoir clearance.

Determinants of species-specific utilization of ACE2 by human and animal coronaviruses.

Utilization of human ACE2 allowed several bat coronaviruses (CoVs), including the causative agent of COVID-19, to infect humans directly or via intermediate hosts. However, the determinants of species-specific differences in ACE2 usage and the frequency of the ability of animal CoVs to use human ACE2 are poorly understood. Here we applied VSV pseudoviruses to analyze the ability of Spike proteins from 26 human or animal CoVs to use ACE2 receptors across nine reservoir, potential intermediate and human hosts. We show that SARS-CoV-2 Omicron variants evolved towards more efficient ACE2 usage but mutation of R493Q in BA.4/5 and XBB Spike proteins disrupts utilization of ACE2 from Greater horseshoe bats. Variations in ACE2 residues 31, 41 and 354 govern species-specific differences in usage by coronaviral Spike proteins. Mutation of T403R allows the RaTG13 bat CoV Spike to efficiently use all ACE2 orthologs for viral entry. Sera from COVID-19 vaccinated individuals neutralize the Spike proteins of various bat Sarbecoviruses. Our results define determinants of ACE2 receptor usage of diverse CoVs and suggest that COVID-19 vaccination may protect against future zoonoses of bat coronaviruses.

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.

Cryptic SARS-CoV-2 lineage identified on two mink farms as a possible result of long-term undetected circulation in an unknown animal reservoir, Poland, November 2022 to January 2023.

In late 2022 and early 2023, SARS-CoV-2 infections were detected on three mink farms in Poland situated within a few km from each other. Whole-genome sequencing of the viruses on two of the farms showed that they were related to a virus identified in humans in the same region 2 years before (B.1.1.307 lineage). Many mutations were found, including in the S protein typical of adaptations to the mink host. The origin of the virus remains to be determined.

Ability of Non-Hosts and Cucurbitaceous Weeds to Transmit Cucumber Green Mottle Mosaic Virus.

Cucumber green mottle mosaic virus (CGMMV) is a Tobamovirus of economic importance affecting cucurbit crops and Asian cucurbit vegetables. Non-host crops of CGMMV, including capsicum (Capsicum annum), sweetcorn (Zea mays), and okra (Abelmoschus esculentus), were tested for their susceptibility to the virus, with field and glasshouse trials undertaken. After 12 weeks post-sowing, the crops were tested for the presence of CGMMV, and in all cases, no CGMMV was detected. Commonly found within the growing regions of cucurbits and melons worldwide are weeds, such as black nightshade (Solanum nigrum), wild gooseberry (Physalis minima), pigweed (Portulaca oleracea), and Amaranth species. Several weeds/grasses were tested for their ability to become infected with CGMMV by inoculating weeds directly with CGMMV and routinely testing over a period of eight weeks. Amaranthus viridis was found to be susceptible, with 50% of the weeds becoming infected with CGMMV. To further analyse this, six Amaranth samples were used as inoculum on four watermelon seedlings per sample and tested after eight weeks. CGMMV was detected in three of six watermelon bulk samples, indicating that A. viridis is a potential host/reservoir for CGMMV. Further research into the relationship between CGMMV and weed hosts is required. This research also highlights the importance of proper weed management to effectively manage CGMMV.

First report on the detection of Japanese encephalitis virus in fruit bats from India.

Japanese encephalitis (JE) is a mosquito borne viral zoonotic disease and JE virus (JEV) is responsible for causing several children deaths every year in India. Since 1978, cases of JE have been reported from Gorakhpur district of Uttar Pradesh state annually. The knowledge on the role played by wildlife reservoirs in the sylvatic transmission and maintenance of JE virus remains limited. Bats are reservoir hosts for several emerging and re-emerging viral pathogens but their role in zoonotic cycle of JEV has not been elucidated yet. In Gorakhpur district of Uttar Pradesh, 52 fruit bats were found dead on 26 May 2020. The post-mortem report of the bat samples conducted at the Indian Veterinary Research Institute stated that the bats died due to brain hemorrhage, caused by excessive heat. The brain tissue samples of the bats were subjected to investigation using molecular techniques to determine the presence of JEV. The present work reports for the first time the detection of JEV in brain samples of bats from India. The viral load ranging from 8 to 18 copies/reaction was detected in brain samples by TaqMan real Time RT-PCR. The low viral load might be the reason for the absence of apparent clinical signs in bats and suggests the probable role of fruit bats in maintaining the JEV in nature.

Monkeypox could establish new reservoirs in animals.

Concern grows that virus will get footholds in species outside Africa.

A Review of Mammarenaviruses and Rodent Reservoirs in the Americas.

In the Americas, infectious viral diseases caused by viruses of the genus Mammarenavirus have been reported since the 1960s. Such diseases have commonly been associated with land use changes, which favor abundance of generalist rodent species. In the Americas-where the rates of land use change are among the highest worldwide-at least 1326 of all 2277 known rodent species have been reported. We conducted a literature review of studies between 1960 and 2020, to establish the current and historical knowledge about genotypes of mammarenaviruses and their rodent reservoirs in the Americas. Our overall goal was to show the importance of focusing research efforts on the American continent, since the conditions exist for future viral hemorrhagic fever (VHF) outbreaks caused by rodent-borne viruses, in turn, carried by widely distributed rodents. We found 47 species identified down to the species level, and one species identified only down to the genus level (Oryzomys sp.), reported in the Americas as reservoirs of mammarenaviruses, most these are ecological generalists. These species associate with 29 genotypes of Mammarenavirus, seven of which have been linked to VHFs in humans. We also highlight the need to monitor these species, in order to prevent viral disease outbreaks in the region.

American alligators are capable of West Nile virus amplification, mosquito infection and transmission.

West Nile virus (WNV) overwintering is poorly understood and likely multifactorial. Interest in alligators as a potential amplifying host arose when it was shown that they develop viremias theoretically sufficient to infect mosquitoes. We examined potential ways in which alligators may contribute to the natural ecology of WNV. We experimentally demonstrated that alligators are capable of WNV amplification with subsequent mosquito infection and transmission capability, that WNV-infected mosquitoes readily infect alligators and that water can serve as a source of infection for alligators but does not easily serve as in intermediate means for transmission between birds and alligators. These findings indicate potential mechanisms for maintenance of WNV outside of the primary bird-mosquito transmission cycle.

Presence of Recombinant Bat Coronavirus GCCDC1 in Cambodian Bats.

Bats have been recognized as an exceptional viral reservoir, especially for coronaviruses. At least three bat zoonotic coronaviruses (SARS-CoV, MERS-CoV and SARS-CoV-2) have been shown to cause severe diseases in humans and it is expected more will emerge. One of the major features of CoVs is that they are all highly prone to recombination. An extreme example is the insertion of the P10 gene from reoviruses in the bat CoV GCCDC1, first discovered in Rousettus leschenaultii bats in China. Here, we report the detection of GCCDC1 in four different bat species (Eonycteris spelaea, Cynopterus sphinx, Rhinolophus shameli and Rousettus sp.) in Cambodia. This finding demonstrates a much broader geographic and bat species range for this virus and indicates common cross-species transmission. Interestingly, one of the bat samples showed a co-infection with an Alpha CoV most closely related to RsYN14, a virus recently discovered in the same genus (Rhinolophus) of bat in Yunnan, China, 2020. Taken together, our latest findings highlight the need to conduct active surveillance in bats to assess the risk of emerging CoVs, especially in Southeast Asia.

Alphacoronaviruses Are Common in Bats in the Upper Midwestern United States.

Bats are a reservoir for coronaviruses (CoVs) that periodically spill over to humans, as evidenced by severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2. A collection of 174 bat samples originating from South Dakota, Minnesota, Iowa, and Nebraska submitted for rabies virus testing due to human exposure were analyzed using a pan-coronavirus PCR. A previously partially characterized CoV, Eptesicus bat CoV, was identified in 12 (6.9%) samples by nested RT-PCR. Six near-complete genomes were determined. Genetic analysis found a high similarity between all CoV-positive samples, Rocky Mountain bat CoV 65 and alphacoronavirus HCQD-2020 recently identified in South Korea. Phylogenetic analysis of genome sequences showed EbCoV is closely related to bat CoV HKU2 and swine acute diarrhea syndrome CoV; however, topological incongruences were noted for the spike gene that was more closely related to porcine epidemic diarrhea virus. Similar to some alphaCoVs, a novel gene, ORF7, was discovered downstream of the nucleocapsid, whose protein lacked similarity to known proteins. The widespread circulation of EbCoV with similarities to bat viruses that have spilled over to swine warrants further surveillance.

Reservoirs of Red-Spotted Grouper Nervous Necrosis Virus (RGNNV) in Squid and Shrimp Species of Northern Alboran Sea.

The production of the aquaculture industry has increased to be equal to that of the world fisheries in recent years. However, aquaculture production faces threats such as infectious diseases. Betanodaviruses induce a neurological disease that affects fish species worldwide and is caused by nervous necrosis virus (NNV). NNV has a nude capsid protecting a bipartite RNA genome that consists of molecules RNA1 and RNA2. Four NNV strains distributed worldwide are discriminated according to sequence homology of the capsid protein encoded by RNA2. Since its first description over 30 years ago, the virus has expanded and reassortant strains have appeared. Preventive treatments prioritize the RGNNV (red-spotted grouper nervous necrosis virus) strain that has the highest optimum temperature for replication and the broadest range of susceptible species. There is strong concern about the spreading of NNV in the mariculture industry through contaminated diet. To surveil natural reservoirs of NNV in the western Mediterranean Sea, we collected invertebrate species in 2015 in the Alboran Sea. We report the detection of the RGNNV strain in two species of cephalopod mollusks (Alloteuthis media and Abralia veranyi), and in one decapod crustacean (Plesionika heterocarpus). According to RNA2 sequences obtained from invertebrate species and reported to date in the Mediterranean Sea, the strain RGNNV is predominant in this semienclosed sea. Neither an ecosystem- nor host-driven distribution of RGNNV were observed in the Mediterranean basin.

Full Genome of batCoV/MinFul/2018/SriLanka, a Novel Alpha-Coronavirus Detected in Miniopterus fuliginosus, Sri Lanka.

Coronaviruses (CoV) are divided into the genera α-CoVs, ß-CoVs, γ-CoVs and δ-CoVs. Of these, α-CoVs and ß-CoVs are solely capable of causing infections in humans, resulting in mild to severe respiratory symptoms. Bats have been identified as natural reservoir hosts for CoVs belonging to these two genera. Consequently, research on bat populations, CoV prevalence in bats and genetic characterization of bat CoVs is of special interest to investigate the potential transmission risks. We present the genome sequence of a novel α-CoV strain detected in rectal swab samples of Miniopterus fuliginosus bats from a colony in the Wavul Galge cave (Koslanda, Sri Lanka). The novel strain is highly similar to Miniopterus bat coronavirus 1, an α-CoV located in the subgenus of Minunacoviruses. Phylogenetic reconstruction revealed a high identity of the novel strain to other α-CoVs derived from Miniopterus bats, while human-pathogenic α-CoV strains like HCoV-229E and HCoV-NL63 were more distantly related. Comparison with selected bat-related and human-pathogenic strains of the ß-CoV genus showed low identities of ~40%. Analyses of the different genes on nucleotide and amino acid level revealed that the non-structural ORF1a/1b are more conserved among α-CoVs and ß-CoVs, while there are higher variations in the structural proteins known to be important for host specificity. The novel strain was named batCoV/MinFul/2018/SriLanka and had a prevalence of 50% (66/130) in rectal swab samples and 58% (61/104) in feces samples that were collected from Miniopterus bats in Wavul Galge cave. Based on the differences between strain batCoV/MinFul/2018/SriLanka and human-pathogenic α-CoVs and ß-CoVs, we conclude that there is a rather low transmission risk to humans. Further studies in the Wavul Galge cave and at other locations in Sri Lanka will give more detailed information about the prevalence of this virus.

Genomic Bootstrap Barcodes and Their Application to Study the Evolution of Sarbecoviruses.

Recombination creates mosaic genomes containing regions with mixed ancestry, and the accumulation of such events over time can complicate greatly many aspects of evolutionary inference. Here, we developed a sliding window bootstrap (SWB) method to generate genomic bootstrap (GB) barcodes to highlight the regions supporting phylogenetic relationships. The method was applied to an alignment of 56 sarbecoviruses, including SARS-CoV and SARS-CoV-2, responsible for the SARS epidemic and COVID-19 pandemic, respectively. The SWB analyses were also used to construct a consensus tree showing the most reliable relationships and better interpret hidden phylogenetic signals. Our results revealed that most relationships were supported by just a few genomic regions and confirmed that three divergent lineages could be found in bats from Yunnan: SCoVrC, which groups SARS-CoV related coronaviruses from China; SCoV2rC, which includes SARS-CoV-2 related coronaviruses from Southeast Asia and Yunnan; and YunSar, which contains a few highly divergent viruses recently described in Yunnan. The GB barcodes showed evidence for ancient recombination between SCoV2rC and YunSar genomes, as well as more recent recombination events between SCoVrC and SCoV2rC genomes. The recombination and phylogeographic patterns suggest a strong host-dependent selection of the viral RNA-dependent RNA polymerase. In addition, SARS-CoV-2 appears as a mosaic genome composed of regions sharing recent ancestry with three bat SCoV2rCs from Yunnan (RmYN02, RpYN06, and RaTG13) or related to more ancient ancestors in bats from Yunnan and Southeast Asia. Finally, our results suggest that viral circular RNAs may be key molecules for the mechanism of recombination.

Multiple spillovers from humans and onward transmission of SARS-CoV-2 in white-tailed deer.

Many animal species are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and could act as reservoirs; however, transmission in free-living animals has not been documented. White-tailed deer, the predominant cervid in North America, are susceptible to SARS-CoV-2 infection, and experimentally infected fawns can transmit the virus. To test the hypothesis that SARS-CoV-2 is circulating in deer, 283 retropharyngeal lymph node (RPLN) samples collected from 151 free-living and 132 captive deer in Iowa from April 2020 through January of 2021 were assayed for the presence of SARS-CoV-2 RNA. Ninety-four of the 283 (33.2%) deer samples were positive for SARS-CoV-2 RNA as assessed by RT-PCR. Notably, following the November 2020 peak of human cases in Iowa, and coinciding with the onset of winter and the peak deer hunting season, SARS-CoV-2 RNA was detected in 80 of 97 (82.5%) RPLN samples collected over a 7-wk period. Whole genome sequencing of all 94 positive RPLN samples identified 12 SARS-CoV-2 lineages, with B.1.2 (n = 51; 54.5%) and B.1.311 (n = 19; 20%) accounting for ∼75% of all samples. The geographic distribution and nesting of clusters of deer and human lineages strongly suggest multiple human-to-deer transmission events followed by subsequent deer-to-deer spread. These discoveries have important implications for the long-term persistence of the SARS-CoV-2 pandemic. Our findings highlight an urgent need for a robust and proactive "One Health" approach to obtain enhanced understanding of the ecology, molecular evolution, and dissemination of SARS-CoV-2.

Molecular identification of Leishmania RNA virus in cutaneous leishmaniasis patients and rodent reservoirs in Isfahan province, Iran.

Leishmania RNA virus (LRV) is a double-strand RNA virus that was first detected in members of the Leishmania viannia in the New World. The present study aimed to investigate the presence of LRV in the Leishmania species isolated from cutaneous leishmaniasis (CL) patients and rodents as reservoirs in Isfahan province an old zoonotic CL focus, center of Iran. Totally, 85 samples were collected from CL patients (n = 80) and rodent reservoirs (n = 5) from different regions of Isfahan province. Species identification was determined using the PCR-RFLP method. Viral dsRNA was extracted and for observation of 5.3 kb dsRNA on an agarose gel. The presence of LRV was surveyed using the Semi-nested PCR method. For phylogenetic analyzes, 6 samples of 13 isolates were sequenced and a phylogenetic tree was drawn by MEGA7 version 7.0.26. Of 80 Leishmania isolates recovered from the patients with CL, 79 and only one were identified as L. major and L. tropica, respectively. Also, the PCR assays detected four L. major and one L. turanica in five assessed Rhombomys opimus as the rodent reservoirs. LRV was detected only in Leishmania species isolated from 13 species of 85 (15.3%) CL including (L. major, n = 12) and (L. tropica, n = 1). Phylogenetic analysis showed that they were belonged to LRV2 and had the highest similarity with Iranian reference LRV2 in GenBank. Our results showed that the LRV2 was present in cutaneous Leishmania species in Isfahan province is the most historical and touristic province of Iran. In the study LRV was not reported from rodent reservoirs, it may be due to the small sample size. Phylogenetic analysis of current sequences demonstrated that these isolates belong to the registered LRV2 of the Old World.

Exploring the Role of Innate Lymphocytes in the Immune System of Bats and Virus-Host Interactions.

Bats are reservoirs of a large number of viruses of global public health significance, including the ancestral virus for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the causative agent of coronavirus disease 2019 (COVID-19). Although bats are natural carriers of multiple pathogenic viruses, they rarely display signs of disease. Recent insights suggest that bats have a more balanced host defense and tolerance system to viral infections that may be linked to the evolutionary adaptation to powered flight. Therefore, a deeper understanding of bat immune system may provide intervention strategies to prevent zoonotic disease transmission and to identify new therapeutic targets. Similar to other eutherian mammals, bats have both innate and adaptive immune systems that have evolved to detect and respond to invading pathogens. Bridging these two systems are innate lymphocytes, which are highly abundant within circulation and barrier tissues. These cells share the characteristics of both innate and adaptive immune cells and are poised to mount rapid effector responses. They are ideally suited as the first line of defense against early stages of viral infections. Here, we will focus on the current knowledge of innate lymphocytes in bats, their function, and their potential role in host-pathogen interactions. Moreover, given that studies into bat immune systems are often hindered by a lack of bat-specific research tools, we will discuss strategies that may aid future research in bat immunity, including the potential use of organoid models to delineate the interplay between innate lymphocytes, bat viruses, and host tolerance.