New Bacteriophages with Podoviridal Morphotypes Active against Yersinia pestis: Characterization and Application Potential.

Phages of highly pathogenic bacteria represent an area of growing interest for bacterial detection and identification and subspecies typing, as well as for phage therapy and environmental decontamination. Eight new phages-YpEc56, YpEc56D, YpEc57, YpEe58, YpEc1, YpEc2, YpEc11, and YpYeO9-expressing lytic activity towards Yersinia pestis revealed a virion morphology consistent with the Podoviridae morphotype. These phages lyse all 68 strains from 2 different sets of Y. pestis isolates, thus limiting their potential application for subtyping of Y. pestis strains but making them rather promising in terms of infection control. Two phages-YpYeO9 and YpEc11-were selected for detailed studies based on their source of isolation and lytic cross activity towards other Enterobacteriaceae. The full genome sequencing demonstrated the virulent nature of new phages. Phage YpYeO9 was identified as a member of the Teseptimavirus genus and YpEc11 was identified as a member of the Helsettvirus genus, thereby representing new species. A bacterial challenge assay in liquid microcosm with a YpYeO9/YpEc11 phage mixture showed elimination of Y. pestis EV76 during 4 h at a P/B ratio of 1000:1. These results, in combination with high lysis stability results of phages in liquid culture, the low frequency of formation of phage resistant mutants, and their viability under different physical-chemical factors indicate their potential for their practical use as an antibacterial mean.

Economic evaluation of the Hepatitis C virus elimination program in the country of Georgia, 2015 to 2017.

BACKGROUND AND AIMS: In 2015, the country of Georgia launched an elimination program aiming to reduce the prevalence of Hepatitis C virus (HCV) infection by 90% from 5.4% prevalence (~150 000 people). During the first 2.5 years of the program, 770 832 people were screened, 48 575 were diagnosed with active HCV infection, and 41 483 patients were treated with direct-acting antiviral (DAA)-based regimens, with a >95% cure rate. METHODS: We modelled the incremental cost-effectiveness ratio (ICER) of HCV screening, diagnosis and treatment between April 2015 and November 2017 compared to no treatment, in terms of cost per quality-adjusted life year (QALY) gained in 2017 US dollars, with a 3% discount rate over 25 years. We compared the ICER to willingness-to-pay (WTP) thresholds of US$4357 (GDP) and US$871 (opportunity cost) per QALY gained. RESULTS: The average cost of screening, HCV viremia testing, and treatment per patient treated was $386 to the provider, $225 to the patient and $1042 for generic DAAs. At 3% discount, 0.57 QALYs were gained per patient treated. The ICER from the perspective of the provider including generic DAAs was $2285 per QALY gained, which is cost-effective at the $4357 WTP threshold, while if patient costs are included, it is just above the threshold at $4398/QALY. All other scenarios examined in sensitivity analyses remain cost-effective except for assuming a shorter time horizon to the end of 2025 or including the list price DAA cost. Reducing or excluding DAA costs reduced the ICER below the opportunity-cost WTP threshold. CONCLUSIONS: The Georgian HCV elimination program provides valuable evidence that national programs for scaling up HCV screening and treatment for achieving HCV elimination can be cost-effective.

Comparison of Antimicrobial Susceptibility Profiles of Thermotolerant Campylobacter spp. Isolated from Human and Poultry Samples in Georgia (Caucasus).

Antimicrobial resistance remains a public health concern globally. This study presents antimicrobial resistance by microdilution and genetic diversity by the whole-genome sequencing of Campylobacter spp. from human and poultry samples isolated in Georgia in 2020/2021. The major species in poultry samples was C. coli, while C. jejuni was preferentially isolated from human samples. Resistance against tetracycline was highest (100%) in C. coli from industrial chicken and lowest in C. jejuni from clinical isolates (36%), while resistance against ciprofloxacin varied from 80% in C. jejuni from backyard chicken to 100% in C. jejuni and C. coli from industrial chicken. The point mutations in gyrA (T86I) and tet (O) genes were detected as resistance determinants for (fluoro-)quinolone or tetracycline resistance, respectively. Ertapenem resistance is still enigmatic. All isolates displayed sensitivity towards erythromycin, gentamicin and chloramphenicol. Multi-resistance was more frequently observed in C. coli than in C. jejuni, irrespective of the isolation matrix, and in chicken isolates compared to human isolates, independent of the Campylobacter species. The Georgian strains showed high variability of multi-locus sequence types (ST), including novel STs. This study provides the first antibiotic resistance data from Campylobacter spp. in Georgia and addresses the need for follow-up monitoring programs.

Campylobacteriosis, Shigellosis and Salmonellosis in Hospitalized Children with Acute Inflammatory Diarrhea in Georgia.

This is the first study on campylobacteriosis carried out in Georgia. It targeted 382 hospitalized children with acute inflammatory diarrhea. The study was conducted between July 2020 to July 2021 based on the main infection clinic of the capital city. Culture-based bacteriological methods were followed by phenotypic and Real-time PCR tests for bacterial confirmation and identification. The data revealed recent epidemiologic prevalences of the three main causative bacteria in the target population. Shigella sonnei with 19.1% (95% CI: 15.2-23.4%) was the most frequently detected pathogen followed by Campylobacter spp. with 12.3% (95% CI: 9.2-16.0%) and Salmonella spp. with 4.9% (95% CI: 3.0-7.6%). However, in 63.6% of the samples, the causative agent remained unknown. Species differentiation of Campylobacter spp. revealed 81% Campylobacter jejuni and 19% Campylobacter coli. An epidemiological pyramid with estimated magnification factors may give more insights into the burden of campylobacteriosis among the studied population, resulting in a putative annual incidence of 6 per 1000 children in Tbilisi. Children with campylobacteriosis were younger (median age 40 months (interquartile range (IQR) 22-95)) than with shigellosis (median age 92 months (interquartile range (IQR) 52-140)). However, no statistically significant difference was found with the age range of patients with campylobacteriosis and salmonellosis as well as with salmonellosis and shigellosis. In conclusion, Campylobacter spp. may be suspected to be the second most frequent bacterial causative agent of acute inflammatory diarrhea in hospitalized children and the primary cause in the 0-3 age group in Georgia. In addition, Campylobacter CROMagar showed better selectivity in comparison to mCCDA selective agar of stool samples in our study.

A Comparison of the Adaptive Immune Response between Recovered Anthrax Patients and Individuals Receiving Three Different Anthrax Vaccines.

Several different human vaccines are available to protect against anthrax. We compared the human adaptive immune responses generated by three different anthrax vaccines or by previous exposure to cutaneous anthrax. Adaptive immunity was measured by ELISPOT to count cells that produce interferon (IFN)-γ in response to restimulation ex vivo with the anthrax toxin components PA, LF and EF and by measuring circulating IgG specific to these antigens. Neutralising activity of antisera against anthrax toxin was also assayed. We found that the different exposures to anthrax antigens promoted varying immune responses. Cutaneous anthrax promoted strong IFN-γ responses to all three antigens and antibody responses to PA and LF. The American AVA and Russian LAAV vaccines induced antibody responses to PA only. The British AVP vaccine produced IFN-γ responses to EF and antibody responses to all three antigens. Anti-PA (in AVA and LAAV vaccinees) or anti-LF (in AVP vaccinees) antibody titres correlated with toxin neutralisation activities. Our study is the first to compare all three vaccines in humans and show the diversity of responses against anthrax antigens.

Expansion of brucellosis detection in the country of Georgia by screening household members of cases and neighboring community members.

BACKGROUND: Brucellosis is considered as endemic zoonotic disease in the country of Georgia. However, the burden of the disease on a household level is not known. Therefore, this study sought to determine the benefits of active surveillance coupled to serological screening for the early detection of brucellosis among close contacts of brucellosis cases. METHODS: We used an active surveillance approach to estimate the rate of seropositivity among household family members and neighboring community members of brucellosis index cases. All participants were screened using the serum tube agglutination test (SAT). Blood cultures were performed, obtained isolates were identified by a bacteriological algorithm, and confirmed as Brucella spp. using real-time PCR. Further confirmation of Brucella species was done using the AMOS PCR assay. RESULTS: A total of 141 participants enrolled. Of these, 27 were brucellosis index cases, 86 were household family members, and 28 were neighboring community members. The serological evidence of brucellosis in the household member group was 7% and the rate at the household level was 21%. No screened community members were Brucella seropositive. Majority of brucellosis cases were caused by B. melitensis; only one index case was linked to B. abortus. CONCLUSION: We found evidence of brucellosis infection among household family members of brucellosis index cases. B. melitensis was the most common species obtained. Findings of this active surveillance study highlight the importance of screening household family members of brucellosis cases and of the use of culture methods to identify Brucella species in the country of 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.

Human cutaneous anthrax, Georgia 2010-2012.

We assessed the occurrence of human cutaneous anthrax in Georgia during 2010--2012 by examining demographic and spatial characteristics of reported cases. Reporting increased substantially, as did clustering of cases near urban centers. Control efforts, including education about anthrax and livestock vaccination, can be directed at areas of high risk.

Evidence of local persistence of human anthrax in the country of georgia associated with environmental and anthropogenic factors.

BACKGROUND: Anthrax is a soil-borne disease caused by the bacterium Bacillus anthracis and is considered a neglected zoonosis. In the country of Georgia, recent reports have indicated an increase in the incidence of human anthrax. Identifying sub-national areas of increased risk may help direct appropriate public health control measures. The purpose of this study was to evaluate the spatial distribution of human anthrax and identify environmental/anthropogenic factors associated with persistent clusters. METHODS/FINDINGS: A database of human cutaneous anthrax in Georgia during the period 2000-2009 was constructed using a geographic information system (GIS) with case data recorded to the community location. The spatial scan statistic was used to identify persistence of human cutaneous anthrax. Risk factors related to clusters of persistence were modeled using a multivariate logistic regression. Areas of persistence were identified in the southeastern part of the country. Results indicated that the persistence of human cutaneous anthrax showed a strong positive association with soil pH and urban areas. CONCLUSIONS/SIGNIFICANCE: Anthrax represents a persistent threat to public and veterinary health in Georgia. The findings here showed that the local level heterogeneity in the persistence of human cutaneous anthrax necessitates directed interventions to mitigate the disease. High risk areas identified in this study can be targeted for public health control measures such as farmer education and livestock vaccination campaigns.

Phylogeography of Francisella tularensis subspecies holarctica from the country of Georgia.

BACKGROUND: Francisella tularensis, the causative agent of tularemia, displays subspecies-specific differences in virulence, geographic distribution, and genetic diversity. F. tularensis subsp. holarctica is widely distributed throughout the Northern Hemisphere. In Europe, F. tularensis subsp. holarctica isolates have largely been assigned to two phylogenetic groups that have specific geographic distributions. Most isolates from Western Europe are assigned to the B.Br.FTNF002-00 group, whereas most isolates from Eastern Europe are assigned to numerous lineages within the B.Br.013 group. The eastern geographic extent of the B.Br.013 group is currently unknown due to a lack of phylogenetic knowledge about populations at the European/Asian juncture and in Asia. In this study, we address this knowledge gap by describing the phylogenetic structure of F. tularensis subsp. holarctica isolates from the country of Georgia, and by placing these isolates into a global phylogeographic context. RESULTS: We identified a new genetic lineage of F. tularensis subsp. holarctica from Georgia that belongs to the B.Br.013 group. This new lineage is genetically and geographically distinct from lineages previously described from the B.Br.013 group from Central-Eastern Europe. Importantly, this new lineage is basal within the B.Br.013 group, indicating the Georgian lineage diverged before the diversification of the other known B.Br.013 lineages. Although two isolates from the Georgian lineage were collected nearby in the Ukrainian region of Crimea, all other global isolates assigned to this lineage were collected in Georgia. This restricted geographic distribution, as well as the high levels of genetic diversity within the lineage, is consistent with a relatively older origin and localized differentiation. CONCLUSIONS: We identified a new lineage of F. tularensis subsp. holarctica from Georgia that appears to have an older origin than any other diversified lineages previously described from the B.Br.013 group. This finding suggests that additional phylogenetic studies of F. tularensis subsp. holarctica populations in Eastern Europe and Asia have the potential to yield important new insights into the evolutionary history and phylogeography of this broadly dispersed F. tularensis subspecies.

Characterization of pPCP1 Plasmids in Yersinia pestis Strains Isolated from the Former Soviet Union.

Complete sequences of 9.5-kb pPCP1 plasmids in three Yersinia pestis strains from the former Soviet Union (FSU) were determined and compared with those of pPCP1 plasmids in three well-characterized, non-FSU Y. pestis strains (KIM, CO92, and 91001). Two of the FSU plasmids were from strains C2614 and C2944, isolated from plague foci in Russia, and one plasmid was from strain C790 from Kyrgyzstan. Sequence analyses identified four sequence types among the six plasmids. The pPCP1 plasmids in the FSU strains were most genetically related to the pPCP1 plasmid in the KIM strain and least related to the pPCP1 plasmid in Y. pestis 91001. The FSU strains generally had larger pPCP1 plasmid copy numbers compared to strain CO92. Expression of the plasmid's pla gene was significantly (P ≤ .05) higher in strain C2944 than in strain CO92. Given pla's role in Y. pestis virulence, this difference may have important implications for the strain's virulence.

Entamoeba histolytica: the serine-rich gene polymorphism-based genetic variability of clinical isolates from Georgia.

It is generally accepted that a majority of individuals infected by Entamoeba histolytica do not develop symptomatic disease. However, the parasite and the host factors contributing to the development of the disease, remain undetermined. It is also unclear why certain individuals develop extra-intestinal amebiasis without exhibiting apparent intestinal symptoms. An outbreak of amebic liver abscess in Tbilisi, Georgia in 1998-1999 suggested that the causative E. histolytica strain had an unusual propensity for extra-intestinal spread. To correlate the genetic differences with pathogenic potential of the parasite, we have examined the SREHP gene polymorphisms among Georgian E. histolytica isolates. Comparison of polymorphic patterns revealed the presence of several different genotypes of E. histolytica, thus preventing an association of a single genotype with hepatic disease, but supporting the previous finding of extensive genetic diversity among E. histolytica isolates from the same geographic origin.