Geographic distribution and genetic characterization of poxviruses from human infections in Georgia, 2009-2014.

Anthrax is endemic in Georgia, as are multiple zoonotic poxviruses. Poxvirus-associated infections share some clinical manifestations and exposure risks with anthrax, and so it is important to distinguish between the two. With this in mind, an archived collection of anthrax-negative DNA samples was retrospectively screened for poxviruses, and of the 148 human samples tested, 64 were positive. Sequence analysis confirmed the presence of orf virus, bovine papular stomatitis virus, and pseudocowpox virus. This study provides evidence of previously unrecognized poxvirus infections in Georgia and highlights the benefit of the timely identification of such infections by improving laboratory capacity.

Human infection with a zoonotic orthopoxvirus in the country of Georgia.

During 2013, cutaneous lesions developed in two men in the country of Georgia after they were exposed to ill cows. The men had never received vaccination against smallpox. Tests of lesion material with the use of a quantitative real-time polymerase-chain-reaction assay for non-variola virus orthopoxviruses were positive, and DNA sequence analysis implicated a novel orthopoxvirus species. During the ensuing epidemiologic investigation, no additional human cases were identified. However, serologic evidence of exposure to an orthopoxvirus was detected in cows in the patients' herd and in captured rodents and shrews. A third case of human infection that occurred in 2010 was diagnosed retrospectively during testing of archived specimens that were originally submitted for tests to detect anthrax. Orthopoxvirus infection should be considered in persons in whom cutaneous lesions develop after contact with animals.

Retrospective Proteomic Analysis of Serum After Akhmeta Virus Infection: New Suspect Case Identification and Insights Into Poxvirus Humoral Immunity.

Serologic cross-reactivity, a hallmark of orthopoxvirus (OPXV) infection, makes species-specific diagnosis of infection difficult. In this study, we used a variola virus proteome microarray to characterize and differentiate antibody responses to nonvaccinia OPXV infections from smallpox vaccination. The profile of 2 case patients infected with newly discovered OPXV, Akhmeta virus, exhibited antibody responses of greater intensity and broader recognition of viral proteins and includes the B21/22 family glycoproteins not encoded by vaccinia virus strains used as vaccines. An additional case of Akhmeta virus, or nonvaccinia OPXV infection, was identified through community surveillance of individuals with no or uncertain history of vaccination and no recent infection. The results demonstrate the utility of microarrays for high-resolution mapping of antibody response to determine the nature of OPXV exposure.

Frameworks for Preventing, Detecting, and Controlling Zoonotic Diseases.

Preventing zoonotic diseases requires coordinated actions by government authorities responsible for human and animal health. Constructing the frameworks needed to foster intersectoral collaboration can be approached in many ways. We highlight 3 examples of approaches to implement zoonotic disease prevention and control programs. The first, rabies control in Ethiopia, was implemented using an umbrella approach: a comprehensive program designed for accelerated impact. The second, a monkeypox program in Democratic Republic of the Congo, was implemented in a stepwise manner, whereby incremental improvements and activities were incorporated into the program. The third approach, a pathogen discovery program, applied in the country of Georgia, was designed to characterize and understand the ecology, epidemiology, and pathogenesis of a new zoonotic pathogen. No one approach is superior, but various factors should be taken into account during design, planning, and implementation.

Evidence of Extensive Circulation of Yersinia enterocolitica in Rodents and Shrews in Natural Habitats from Retrospective and Perspective Studies in South Caucasus.

Yersinia enterocolitica culture-positive rodents and shrews were reported in different territories across Georgia during 14 of 17 years of investigations conducted for the period of 1981-1997. In total, Y. enterocolitica was isolated from 2052 rodents (15 species) and 33 shrews. Most isolates were obtained from Microtus arvalis, Rattus norvegicus, Mus musculus, and Apodemus spp. During the prospective study (2017-2019), isolates of Yersinia-like bacteria were cultured from 53 rodents collected in four parts of Georgia. All the Yersinia-like isolates were confirmed as Y. enterocolitica based on the API 20E and the BD Phenix50 tests. Whole-genome (WG) sequencing of five rodents and one shrew strain of Y. enterocolitica revealed that they possessed a set of virulence genes characteristic of the potentially pathogenic strains of biogroup 1A. All isolates lacked distinguished virulence determinants for YstA, Ail, TccC, VirF, and virulence plasmid pYV but carried the genes for YstB, YmoA, HemPR-HmuVSTU, YaxAB, PhlA, PldA, ArsCBR, and a flagellar apparatus. One strain contained a gene highly homologous to heat-labile enterotoxin, a chain of E. coli, a function not previously described for Y. enterocolitica. The WG single-nucleotide polymorphism-based typing placed the isolates in four distinct phylogenetic clusters.

Identification and distribution of nine tick-borne spotted fever group Rickettsiae in the Country of Georgia.

Rickettsial pathogens cause diseases that vary in severity and clinical presentation. Rickettsia species transmitted by ticks are mostly classified within the spotted fever group of rickettsiae (SFGR) and are often associated with febrile diseases. Preliminary studies have detected three human-pathogenic SFGR from ticks in Georgia: Rickettsia aeschlimannii, Rickettsia raoultii, and Rickettsia slovaca. To more broadly assess the presence of tick-borne rickettsiae from Georgia we examined 1594 ticks, representing 18 species from five genera (Ixodes, Hyalomma, Haemaphysalis, Dermacentor, and Rhipicephalus), collected from eight regions of Georgia. A total of 498 tick DNA samples extracted from single ticks or pooled ticks were assessed by molecular methods. Genus-specific Rick17b and species-specific qPCR assays were used to identify six rickettsiae: R. aeschlimannii, R. raoultii, R. slovaca, Rickettsia conorii subsp. conorii, Rickettsia massiliae, and Rickettsia monacensis. Tick samples that were positive for Rickettsia, but not identified by the species-specific assays, were further evaluated by multi-locus sequence typing (MLST) using sequences of four protein-coding genes (gltA, ompA,ompB, sca4). Three additional Rickettsia species were identified by MLST: Candidatus Rickettsia barbariae, Rickettsia helvetica, and Rickettsia hoogstraalii. Overall, nine species of Rickettsia (six human pathogens and three species with unknown pathogenicity) were detected from 12 tick species of five different genera. A distribution map for the tick-borne rickettsiae revealed six newly identified endemic regions in Georgia.

Identification of a Novel Yersinia enterocolitica Strain from Bats in Association with a Bat Die-Off That Occurred in Georgia (Caucasus).

Yersinia entercolitica is a bacterial species within the genus Yersinia, mostly known as a human enteric pathogen, but also recognized as a zoonotic agent widespread in domestic pigs. Findings of this bacterium in wild animals are very limited. The current report presents results of the identification of cultures of Y. entercolitica from dead bats after a massive bat die-off in a cave in western Georgia. The growth of bacterial colonies morphologically suspected as Yersinia was observed from three intestine tissues of 11 bats belonging to the Miniopterus schreibersii species. These three isolates were identified as Y. enterocolitica based on the API29 assay. No growth of Brucella or Francisella bacteria was observed from tissues of dead bats. Full genomes (a size between 4.6-4.7 Mbp) of the Yersinia strains isolated from bats were analyzed. The phylogenetic sequence analyses of the genomes demonstrated that all strains were nearly identical and formed a distinct cluster with the closest similarity to the environmental isolate O:36/1A. The bat isolates represent low-pathogenicity Biotype 1A strains lacking the genes for the Ail, Yst-a, Ysa, and virulence plasmid pYV, while containing the genes for Inv, YstB, and MyfA. Further characterization of the novel strains cultured from bats can provide a clue for the determination of the pathogenic properties of those strains.

Parapoxvirus Infections in the Country of Georgia: A Case Series.

Infections caused by viruses of the parapoxvirus (PPV) genus, including orf and pseudocowpox viruses, are frequently seen in both humans and animals in many regions of the world. These infections are often misdiagnosed or neglected because of the lack of clinician awareness, inadequate diagnostic capacity, and their relatively mild disease presentation, which may result in affected individuals not seeking medical attention. Although PPV infections should be routinely considered in patients with cutaneous lesions, especially in those who have occupational exposure to farm animals, they are often excluded from the differential diagnosis because they are not perceived as serious, resulting in underestimation of the burden of disease. Since 2014, significant enhancements to Georgia's epidemiologic and laboratory capacity have made PPV surveillance and detection possible. In this study, we present information on 27 confirmed cases of PPV infection reported to Georgia's national surveillance system from January 2016 through January 2017.

Genome Sequences of Akhmeta Virus, an Early Divergent Old World Orthopoxvirus.

Annotated whole genome sequences of three isolates of the Akhmeta virus (AKMV), a novel species of orthopoxvirus (OPXV), isolated from the Akhmeta and Vani regions of the country Georgia, are presented and discussed. The AKMV genome is similar in genomic content and structure to that of the cowpox virus (CPXV), but a lower sequence identity was found between AKMV and Old World OPXVs than between other known species of Old World OPXVs. Phylogenetic analysis showed that AKMV diverged prior to other Old World OPXV. AKMV isolates formed a monophyletic clade in the OPXV phylogeny, yet the sequence variability between AKMV isolates was higher than between the monkeypox virus strains in the Congo basin and West Africa. An AKMV isolate from Vani contained approximately six kb sequence in the left terminal region that shared a higher similarity with CPXV than with other AKMV isolates, whereas the rest of the genome was most similar to AKMV, suggesting recombination between AKMV and CPXV in a region containing several host range and virulence genes.

Ten Genome Sequences of Human and Livestock Isolates of Bacillus anthracis from the Country of Georgia.

Bacillus anthracis causes the acute fatal disease anthrax, is a proven biological weapon, and is endemic in Georgia, where human and animal cases are reported annually. Here, we present whole-genome sequences of 10 historical B. anthracis strains from Georgia.

Phylogeography of Bacillus anthracis in the country of Georgia shows evidence of population structuring and is dissimilar to other regional genotypes.

Sequence analyses and subtyping of Bacillus anthracis strains from Georgia reveal a single distinct lineage (Aust94) that is ecologically established. Phylogeographic analysis and comparisons to a global collection reveals a clade that is mostly restricted to Georgia. Within this clade, many groups are found around the country, however at least one subclade is only found in the eastern part. This pattern suggests that dispersal into and out of Georgia has been rare and despite historical dispersion within the country, for at least for one lineage, current spread is limited.