Genomic epidemiology of Mycobacterium bovis infection in sympatric badger and cattle populations in Northern Ireland.

Bovine tuberculosis (bTB) is a costly, epidemiologically complex, multi-host, endemic disease. Lack of understanding of transmission dynamics may undermine eradication efforts. Pathogen whole-genome sequencing improves epidemiological inferences, providing a means to determine the relative importance of inter- and intra-species host transmission for disease persistence. We sequenced an exceptional data set of 619 Mycobacterium bovis isolates from badgers and cattle in a 100 km2 bTB 'hotspot' in Northern Ireland. Historical molecular subtyping data permitted the targeting of an endemic pathogen lineage, whose long-term persistence provided a unique opportunity to study disease transmission dynamics in unparalleled detail. Additionally, to assess whether badger population genetic structure was associated with the spatial distribution of pathogen genetic diversity, we microsatellite genotyped hair samples from 769 badgers trapped in this area. Birth death models and TransPhylo analyses indicated that cattle were likely driving the local epidemic, with transmission from cattle to badgers being more common than badger to cattle. Furthermore, the presence of significant badger population genetic structure in the landscape was not associated with the spatial distribution of M. bovis genetic diversity, suggesting that badger-to-badger transmission is not playing a major role in transmission dynamics. Our data were consistent with badgers playing a smaller role in transmission of M. bovis infection in this study site, compared to cattle. We hypothesize, however, that this minor role may still be important for persistence. Comparison to other areas suggests that M. bovis transmission dynamics are likely to be context dependent, with the role of wildlife being difficult to generalize.

Nanoisland SERS-Substrates for Specific Detection and Quantification of Influenza A Virus.

Surface-enhanced Raman spectroscopy (SERS)-based aptasensors for virus determination have attracted a lot of interest recently. This approach provides both specificity due to an aptamer component and a low limit of detection due to signal enhancement by a SERS substrate. The most successful SERS-based aptasensors have a limit of detection (LoD) of 10-100 viral particles per mL (VP/mL) that is advantageous compared to polymerase chain reactions. These characteristics of the sensors require the use of complex substrates. Previously, we described silver nanoisland SERS-substrate with a reproducible and uniform surface, demonstrating high potency for industrial production and a suboptimal LoD of 4 × 105 VP/mL of influenza A virus. Here we describe a study of the sensor morphology, revealing an unexpected mechanism of signal enhancement through the distortion of the nanoisland layer. A novel modification of the aptasensor was proposed with chromium-enhanced adhesion of silver nanoparticles to the surface as well as elimination of the buffer-dependent distortion-triggering steps. As a result, the LoD of the Influenza A virus was decreased to 190 VP/mL, placing the nanoisland SERS-based aptasensors in the rank of the most powerful sensors for viral detection.

Lithographic SERS Aptasensor for Ultrasensitive Detection of SARS-CoV-2 in Biological Fluids.

In this paper, we propose a technology for the rapid and sensitive detection of the whole viral particles of SARS-CoV-2 using double-labeled DNA aptamers as recognition elements together with the SERS method for detecting the optical response. We report on the development of a SERS-aptasensor based on a reproducible lithographic SERS substrate, featuring the combination of high speed, specificity, and ultrasensitive quantitative detection of SARS-CoV-2 virions. The sensor makes it possible to identify SARS-CoV-2 in very low concentrations (the limit of detection was 100 copies/mL), demonstrating a sensitivity level comparable to the existing diagnostic golden standard-the reverse transcription polymerase chain reaction.

A Combination of Membrane Filtration and Raman-Active DNA Ligand Greatly Enhances Sensitivity of SERS-Based Aptasensors for Influenza A Virus.

Biosensors combining the ultrahigh sensitivity of surface-enhanced Raman scattering (SERS) and the specificity of nucleic acid aptamers have recently drawn attention in the detection of respiratory viruses. The most sensitive SERS-based aptasensors allow determining as low as 104 virus particles per mL that is 100-fold lower than any antibody-based lateral flow tests but 10-100-times higher than a routine polymerase chain reaction with reversed transcription (RT-PCR). Sensitivity of RT-PCR has not been achieved in SERS-based aptasensors despite the usage of sophisticated SERS-active substrates. Here, we proposed a novel design of a SERS-based aptasensor with the limit of detection of just 103 particles per ml of the influenza A virus that approaches closely to RT-PCR sensitivity. The sensor utilizes silver nanoparticles with the simplest preparation instead of sophisticated SERS-active surfaces. The analytical signal is provided by a unique Raman-active dye that competes with the virus for the binding to the G-quadruplex core of the aptamer. The aptasensor functions even with aliquots of the biological fluids due to separation of the off-target molecules by pre-filtration through a polymeric membrane. The aptasensor detects influenza viruses in the range of 1·103-5·1010 virus particles per ml.

High resolution imaging of viruses: Scanning probe microscopy and related techniques.

Scanning probe microscopy is a group of measurements that provides 3D visualization of viruses in different environmental conditions including liquids and air. Besides 3D topography it is possible to measure the properties like mechanical rigidity and stability, adhesion, tendency to crystallization, surface charge, etc. Choosing the right substrate and scanning parameters makes it much easier to obtain reliable data. Rational interpretation of experimental results should take into account possible artifacts, proper filtering and data presentation using specially designed software packages. Animal and human virus characterization is in the focus of many intensive studies because of their potential harm to higher organisms. The article focuses on high-resolution visualization of plant viruses. Tobacco mosaic virus, potato viruses X and B and others are not dangerous for the human being and are widely used in different applications such as vaccine preparation, construction of building units in nanotechnology and material science applications, nanoparticle production and delivery, and even metrology. The methods of virus's deposition, visualization, and consequent image processing and interpretation are described in details. Specific examples of viruses imaging are illustrated using the FemtoScan Online software, which has typical and all the necessary built-in functions for constructing three-dimensional images, their processing and analysis. Despite visible progress in visualizing the viruses using probe microscopy, many unresolved problems still remain. At present time the probe microscopy data on viruses is not systemized. There is no descriptive atlas of the images and morphology as revealed by this type of high resolution microscopy. It is worth emphasizing that new virus investigation methods will appear due to the progress of science.

"Chasing hope": Parents' perspectives on complementary and alternative interventions for children with autism in Kazakhstan.

LAY ABSTRACT: The article reports the findings of a qualitative research study on how and why parents of autistic children in Kazakhstan utilize complementary and alternative medicine. We found that parents turn to complementary and alternative medicine because of the lack of professional care options available to them and in pursuit for hope and opportunities for their children with ASD.

Implementation of a Regional Training Program on African Swine Fever As Part of the Cooperative Biological Engagement Program across the Caucasus Region.

A training and outreach program to increase public awareness of African swine fever (ASF) was implemented by Defense Threat Reduction Agency and the Ministries of Agriculture in Armenia, Georgia, Kazakhstan, and Ukraine. The implementing agency was the company SAFOSO (Switzerland). Integration of this regional effort was administered by subject matter experts for each country. The main teaching effort of this project was to develop a comprehensive regional public outreach campaign through a network of expertise and knowledge for the control and prevention of ASF in four neighboring countries that experience similar issues with this disease. Gaps in disease knowledge, legislation, and outbreak preparedness in each country were all addressed. Because ASF is a pathogen with bioterrorism potential and of great veterinary health importance that is responsible for major economic instability, the project team developed public outreach programs to train veterinarians in the partner countries to accurately and rapidly identify ASF activity and report it to international veterinary health agencies. The project implementers facilitated four regional meetings to develop this outreach program, which was later disseminated in each partner country. Partner country participants were trained as trainers to implement the outreach program in their respective countries. In this paper, we describe the development, execution, and evaluation of the ASF training and outreach program that reached more than 13,000 veterinarians, farmers, and hunters in the partner countries. Additionally, more than 120,000 booklets, flyers, leaflets, guidelines, and posters were distributed during the outreach campaign. Pre- and post-ASF knowledge exams were developed. The overall success of the project was demonstrated in that the principles of developing and conducting a public outreach program were established, and these foundational teachings can be applied within a single country or expanded regionally to disseminate disease information across borders; overall, this method can be modified to raise awareness about many other diseases.

Epidemiology of Brucellosis and Genetic Diversity of Brucella abortus in Kazakhstan.

Brucellosis is a major zoonotic infection in Kazakhstan. However, there is limited data on its incidence in humans and animals, and the genetic diversity of prevalent strains is virtually unstudied. Additionally, there is no detailed overview of Kazakhstan brucellosis control and eradication programs. Here, we analyzed brucellosis epidemiological data, and assessed the effectiveness of eradication strategies employed over the past 70 years to counteract this infection. We also conducted multiple loci variable-number tandem repeat analysis (MLVA) of Brucella abortus strains found in Kazakhstan. We analyzed official data on the incidence of animal brucellosis in Kazakhstan. The records span more than 70 years of anti-brucellosis campaigns, and contain a brief description of the applied control strategies, their effectiveness, and their impact on the incidence in humans. The MLVA-16 method was used to type 94 strains of B. abortus and serial passages of B. abortus 82, a strain used in vaccines. MLVA-8 and MLVA-11 analyses clustered strains into a total of four and seven genotypes, respectively; it is the first time that four of these genotypes have been described. MLVA-16 analysis divided strains into 28 distinct genotypes having genetic similarity coefficient that varies from 60 to100% and a Hunter & Gaston diversity index of 0.871. MST analysis reconstruction revealed clustering into "Kazakhstani-Chinese (Central Asian)", "European" and "American" lines. Detection of multiple genotypes in a single outbreak confirms that poorly controlled trade of livestock plays a crucial role in the spread of infection. Notably, the MLVA-16 profile of the B. abortus 82 strain was unique and did not change during 33 serial passages. MLVA genotyping may thus be useful for epidemiological monitoring of brucellosis, and for tracking the source(s) of infection. We suggest that countrywide application of MLVA genotyping would improve the control of brucellosis in Kazakhstan.

Genetic diversity of Brucella abortus and Brucella melitensis in Kazakhstan using MLVA-16.

Brucellosis is an endemic disease in Central Asia characterized by high infection rates in humans and animals. Currently, little is known about the genetic diversity of Brucella spp. circulating in the region, despite the high prevalence of brucellosis. This study aimed to analyze the genetic diversity of Brucella melitensis and Brucella abortus strains circulating in the Republic of Kazakhstan. We genotyped 128 B. melitensis and 124 B. abortus strains collected in regions with the highest prevalence of brucellosis. Genotyping was performed using multi-locus variable-number tandem-repeat analysis (MLVA). Analysis of a subset of 8 loci (MLVA-8) of 128 B. melitensis strains identified genotypes 42 (n=108), 43 (n=2), and 63 (n=19) related to the 'East Mediterranean' group. An MLVA-16 assay sorted 128 B. melitensis strains into 25 different genotypes. Excluding one variable locus, MLVA-15 of B. melitensis was distinct from strains originating in the Mediterranean region; however, 77% of them were identical to strains isolated in China. A minimum spanning tree for B. melitensis using MLVA-15 analysis clustered the local strains together with strains previously collected in China. MLVA-8 analysis of 124 B. abortus strains identified them as genotype 36, suggesting Eurasian distribution of this lineage. Complete MLVA-16 assay analysis clustered the strains into five genotypes, revealing little diversity of B. abortus when compared on the global scale. A minimum spanning tree for B. abortus obtained using MLVA-15 analysis clustered the 2 most prevalent genotypes (n=117) together with strains previously collected in China. Thus, MLVA analysis was used to characterize 252 strains of Brucella collected in Kazakhstan. The analysis revealed genetic homogeneity among the strains. Interestingly, identical MLVA-15 profiles were found in seemingly unrelated outbreaks in China, Turkey, and Kazakhstan. Further analysis is needed for better understanding of the epidemiology of brucellosis in Asia.