United States Army Medical Research Directorate - Georgia

We analysed both pooled and individual tick samples collected from four countries in Eastern Europe and the Black Sea region, using metagenome-based nanopore sequencing (NS) and targeted amplification. Initially, 1337 ticks, belonging to 11 species, were screened in 217 pools. Viruses (21 taxa) and human pathogens were detected in 46.5% and 7.3%, respectively. Tick-borne viral pathogens comprised Tacheng Tick Virus 2 (TTV2, 5.9%), Jingmen Tick Virus (JMTV, 0.9%) and Tacheng Tick Virus 1 (TTV1, 0.4%). An association of tick species with individual virus taxa was observed, with the exception of TTV2, which was observed in both Dermacentor and Haemaphysalis species. Individual ticks from pools with pathogen detection were then further screened by targeted amplification and then NS, which provided extensive genome data and revealed probable pathogen Haseki Tick Virus (HTV, 10.2%). Two distinct TTV2 clades were observed in phylogenetic analysis, one of which included closely related Dermacentor reticulatus Uukuviruses. JMTV detection indicated integrated virus sequences. Overall, we observed an expansion of newly documented pathogenic tick-borne viruses into Europe, with TTV1 being identified on the continent for the first time. These viruses should be included in the diagnostic assessment of symptomatic cases associated with tick bites and vector surveillance efforts. NS is shown as a useful tool for monitoring tick-associated pathogens in pooled or individual samples.

Introduction: We evaluated metagenomic nanopore sequencing (NS) in field-collected ticks and compared findings from amplification-based assays. Methods: Forty tick pools collected in Anatolia, Turkey and screened by broad-range or nested polymerase chain reaction (PCR) for Crimean-Congo Hemorrhagic Fever Virus (CCHFV) and Jingmen tick virus (JMTV) were subjected to NS using a standard, cDNA-based metagenome approach. Results: Eleven viruses from seven genera/species were identified. Miviruses Bole tick virus 3 and Xinjiang mivirus 1 were detected in 82.5 and 2.5% of the pools, respectively. Tick phleboviruses were present in 60% of the pools, with four distinct viral variants. JMTV was identified in 60% of the pools, where only 22.5% were PCR-positive. CCHFV sequences characterized as Aigai virus were detected in 50%, where only 15% were detected by PCR. NS produced a statistically significant increase in detection of these viruses. No correlation of total virus, specific virus, or targeted segment read counts was observed between PCR-positive and PCR-negative samples. NS further enabled the initial description of Quaranjavirus sequences in ticks, where human and avian pathogenicity of particular isolates had been previously documented. Discussion: NS was observed to surpass broad-range and nested amplification in detection and to generate sufficient genome-wide data for investigating virus diversity. It can be employed for monitoring pathogens in tick vectors or human/animal clinical samples in hot-spot regions for examining zoonotic spillover.

Accurate identification of tick-borne bacteria, including those associated with rickettsioses, pose significant challenges due to the polymicrobial and polyvectoral nature of the infections. We aimed to carry out a comparative evaluation of a non-targeted metagenomic approach by nanopore sequencing (NS) and commonly used PCR assays amplifying Rickettsia genes in field-collected ticks. The study included a total of 310 ticks, originating from Poland (44.2 %) and Bulgaria (55.8 %). Samples comprised 7 species, the majority of which were Ixodes ricinus (62.9 %), followed by Dermacentor reticulatus (21.2 %). Screening was carried out in 55 pools, using total nucleic acid extractions from individual ticks. NS and ompA/gltA PCRs identified Rickettsia species in 47.3 % and 54.5 % of the pools, respectively. The most frequently detected species were Rickettsia asiatica (27.2 %) and Rickettsia raoultii (21.8 %), followed by Rickettsia monacensis (3.6 %), Rickettsia helvetica (1.8 %), Rickettsia massiliae (1.8 %) and Rickettsia tillamookensis (1.8 %). Phylogeny construction on mutS, uvrD, argS and virB4 sequences and a follow-up deep sequencing further supported R. asiatica identification, documented in Europe for the first time. NS further enabled detection of Anaplasma phagocytophilum (9.1 %), Coxiella burnetii (5.4 %) and Neoehrlichia mikurensis (1.8 %), as well as various endosymbionts of Rickettsia and Coxiella. Co-detection of multiple rickettsial and non-rickettsial bacteria were observed in 16.4 % of the pools with chromosome and plasmid-based contigs. In conclusion, non-targeted metagenomic sequencing was documented as a robust strategy capable of providing a broader view of the tick-borne bacterial pathogen spectrum.

BACKGROUND: Acute diarrhea (AD) can have significant impacts on military troop readiness. Medical providers must understand current trends of enteropathogen antimicrobial resistance (AMR) in service members (SMs) to inform proper, timely treatment options. However, little is known of enteric pathogen profiles across the Military Health System (MHS). The primary objectives of this study were to identify gaps in enteric pathogen surveillance within the MHS, describe the epidemiology of AMR in enteric pathogens, and identify trends across the MHS both within the Continental United States (CONUS) and outside of the Continental United States (OCONUS). METHODS: Health Level 7 (HL7)-formatted laboratory data were queried for all specimens where Salmonella, Shigella, and Campylobacter species, as well as Shiga toxin-producing Escherichia coli (E. coli) (STEC) were isolated and certified between 1 January 2009 - 31 December 2019. Antibiotic susceptibility testing (AST) results were queried and summarized where available. Descriptive statistics were calculated for each organism by specimen source, year, and susceptibility testing availability. RESULTS: Among a total of 13,852 enteric bacterial isolates, 11,877 (86%) were submitted from CONUS locations. Out of 1479 Shigella spp. and 6755 Salmonella spp. isolates, 1221 (83%) and 5019 (74%), respectively, reported any susceptibility results through the MHS. Overall, only 15% of STEC and 4% of Campylobacter spp. specimens had AST results available. Comparing AST reporting at CONUS versus OCONUS locations, AST was reported for 1175 (83%) and 46 (78%) of Shigella isolates at CONUS and OCONUS locations, respectively, and for 4591 (76%) and 428 (63%) of Salmonella isolates at CONUS and OCONUS locations, respectively. CONCLUSIONS: This study revealed inconsistent enteropathogen AST conducted across the MHS, with differing trends between CONUS and OCONUS locations. Additional work is needed to assess pathogen-specific gaps in testing and reporting to develop optimal surveillance that supports the health of the force.

SHP-2 (encoded by PTPN11) is a ubiquitously expressed protein tyrosine phosphatase required for signal transduction by multiple different cell surface receptors. Humans with carry germline SHP-2 mutations develop Noonan syndrome or LEOPARD syndrome, which are characterized by cardiovascular, neurological and skeletal abnormalities.Shp2 is an important signaling agent for growth factors and cytokines. To investigate the Ptpn11 in postnatal myogenesis of mice  we analyzed using immunohistochemistry stem cell markers: Pax 7, Myo D, Myf 5 and Myo G in muscle cells isolated from wild type and Shp2 knockout mice. We used a conditional SHP-2 mouse mutant in which loss of expression of SHP-2 was induced in multiple tissues in response to drug administration. Our findings indicate that all these markers gradually decreased from birth to day 14 in mouse muscle cells. Here we show that Shp2, an intracellular tyrosine phosphatase with two SH2 domains, plays a critical role in mouse muscle development after birth. Our data demonstrate a molecular difference in the control and Sh2 knockout  postnatal myogenic stem cells, and assign to Ptpn11 signaling a key function in satellite cell activity. These findings illustrate an essential role for Shp-2 in muscle growth and remodeling in adults, and reveal some of the cellular and molecular mechanisms involved. The model is predicted to be of further use in understanding how Shp-2 regulates muscle morphogenesis, which could lead to the development of novel therapies for the treatment of malformations in human patients with SHP-2 mutations.

The Dynamic Tumor Microenvironment Theory (DTMT) constitutes a pivotal milestone in tumor research and biochemistry, ushering in a paradigm characterized by substantial depth and comprehensiveness. This theory represents a substantial leap in comprehending the intricacies governing the dynamic interplay between tumor cells and their microenvironment. Through the integration of insights from tumor biology, biochemistry, and systems biology, DTMT provides an all-encompassing perspective, revealing the intricate complexity inherent in the dynamic nature of tumors. This holistic viewpoint not only elucidates the nuanced intricacies of tumor behavior but also catalyzes the development of innovative therapeutic strategies that surpass conventional approaches. The expansive scope of DTMT transcends disciplinary boundaries, important for emerging treatment modalities and advancing personalized medicine. Navigating the intricate network of molecular interactions and systemic influences, DTMT functions as a guiding framework, offering insights beyond the traditional confines of tumor research. DTMT extends to reshaping the landscape of personalized medicine in cancer treatment. The theory's comprehensive understanding of the tumor microenvironment facilitates the identification of novel biomarkers and therapeutic targets, fostering a nuanced and tailored approach to patient care. This expanded view not only enhances our comprehension of intricate interrelationships between diverse biological components but also heralds a new era in precision medicine, where treatment strategies are finely tuned to the unique characteristics of individual tumors. In conclusion, the DTMT transcends conventional boundaries, offering an expansive and intricate exploration of the multifaceted dimensions of tumor research and biochemistry. This all-encompassing framework not only advances our understanding of tumors but also propels the field towards innovative therapeutic frontiers, ultimately redefining the trajectory of cancer research and personalized medicine.

 The Dynamic Cellular Equilibrium Theory of Aging introduces a comprehensive framework to comprehend the complex mechanisms governing the aging process. This theory posits that aging results from the disruption of delicate balance between cellular upkeep mechanisms and the accrual of cellular damage, all regulated by an interplay of genetic, epigenetic, environmental, and stochastic factors. Within this article, an in-depth exploration of the theory is conducted, encompassing diverse aspects such as the dynamics of cellular maintenance, the intricacies of damage accumulation, the sway of genetic and epigenetic forces, the influence exerted by environmental and lifestyle elements, the stochastic characteristics characterizing aging, along with the cellular adaptive retorts to these influences. By drawing upon pertinent scientific literature, this theory not only provides profound insights into the aging process but also furnishes valuable implications for interventions that strive to cultivate healthy aging and protract the human lifespan. The profound impact of genetic and epigenetic influences on the aging is discussed, the theory unraveling the importance of genes and epigenetic marks that choreograph the symphony of aging. Moreover, there is considered the pervasive role of environmental factors, encompassing lifestyle choices, diet, and exposure to toxins, is expounded upon, underscoring their potent role in shaping the aging process. Incorporating the stochastic element of chance events into the narrative of aging, this theory acknowledges the role of random occurrences in the gradual unfolding of cellular degeneration. Furthermore, the remarkable resilience of cells, reflected through adaptive responses, is elucidated, demonstrating the remarkable plasticity cells exhibit in the face of various aging-related challenges.