Characterization of a putative orexin receptor in Ciona intestinalis sheds light on the evolution of the orexin/hypocretin system in chordates.

Tunicates are evolutionary model organisms bridging the gap between vertebrates and invertebrates. A genomic sequence in Ciona intestinalis (CiOX) shows high similarity to vertebrate orexin receptors and protostome allatotropin receptors (ATR). Here, molecular phylogeny suggested that CiOX is divergent from ATRs and human orexin receptors (hOX1/2). However, CiOX appears closer to hOX1/2 than to ATR both in terms of sequence percent identity and in its modelled binding cavity, as suggested by molecular modelling. CiOX was heterologously expressed in a recombinant HEK293 cell system. Human orexins weakly but concentration-dependently activated its Gq signalling (Ca2+ elevation), and the responses were inhibited by the non-selective orexin receptor antagonists TCS 1102 and almorexant, but only weakly by the OX1-selective antagonist SB-334867. Furthermore, the 5-/6-carboxytetramethylrhodamine (TAMRA)-labelled human orexin-A was able to bind to CiOX. Database mining was used to predict a potential endogenous C. intestinalis orexin peptide (Ci-orexin-A). Ci-orexin-A was able to displace TAMRA-orexin-A, but not to induce any calcium response at the CiOX. Consequently, we suggested that the orexin signalling system is conserved in Ciona intestinalis, although the relevant peptide-receptor interaction was not fully elucidated.

Isolation and Characterization of a Novel Aeromonas salmonicida-Infecting Studiervirinae Bacteriophage, JELG-KS1.

Representatives of the bacterial genus Aeromonas are some of the most notorious aquaculture pathogens associated with a range of diseases in different fish species. As the world forges toward the post-antibiotic era, alternative options for combating bacterial pathogens are needed. One such alternative option is phage biocontrol. In this study, a novel podophage-JELG-KS1-infecting Aeromonas salmonicida was retrieved from wastewater along with its host strain. The genome of the JELG-KS1 phage is a 40,505 bp dsDNA molecule with a GC% of 53.42% and 185 bp direct terminal repeats and encodes 53 predicted proteins. Genomic analysis indicates that JELG-KS1 might represent a novel genus within the subfamily Studiervirinae. Podophage JELG-KS1 is a strictly lytic phage without any identifiable virulence or AMR genes that quickly adsorbs onto the surface of host cells to initiate a 48 min long infectious cycle, resulting in the release of 71 ± 12 JELG-KS1 progeny virions per infected cell. JELG-KS1 effectively lyses its host population in vitro, even at very low multiplicities of infection. However, when challenged against a panel of Aeromonas spp. strains associated with diseases in aquaculture, JELG-KS1 shows host-specificity that is confined only to its isolation strain, immediately compromising its potential for Aeromonas spp. biocontrol in aquaculture.

Impact of Anthropogenic Activities on Microbial Community Structure in Riverbed Sediments of East Kazakhstan.

Heavy metal (HMe) pollution in regions with mining and metallurgy activities is known to be a serious environmental problem worldwide. Hydrological processes contribute to the dissemination of HMes (drainage, precipitation, flow rate). The aim of the present study is to investigate the microbial community structure in ten river sediments sampled in different regions of East Kazakhstan, which are contaminated with HMes. The overall degree of sediment contamination with HMes (Cr, Cu, Zn, Pb, and Cd) was assessed using the pollution index Zc, which ranged from 0.43 to 21.6, with the highest in Ridder City (Zc = 21.6) and Ust-Kamenogorsk City, 0.8 km below the dam of the hydroelectric power station (Zc = 19.6). The tested samples considerably differed in organic matter, total carbon, nitrogen, and phosphorus content, as well as in the abundance of HMe-related functional gene families and antibiotic resistance genes. Metagenomic analysis of benthic microorganisms showed the prevalence of Proteobacteria (88.84-97.61%) and Actinobacteria (1.21-5.98%) at the phylum level in all samples. At the class level, Actinobacteria (21.68-57.48%), Betaproteobacteria (19.38-41.17%), and Alphaproteobacteria (10.0-39.78%) were the most common among the classified reads. To the best of our knowledge, this is the first study on the metagenomic characteristics of benthic microbial communities exposed to chronic HMe pressure in different regions of East Kazakhstan.

Prevalence of SARS-CoV-2 in domestic cats (Felis catus) during COVID-19 pandemic in Latvia.

BACKGROUND: The causative agent of the COVID-19 pandemic, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is of zoonotic origin and has shown reverse zoonotic transmissibility. OBJECTIVES: The aim of this cross-sectional study was to investigate the serological and molecular prevalence of SARS-CoV-2 infection in the domestic cat (Felis catus) population from Latvia in natural conditions and subsequently perform viral genome analysis. METHODS: Oropharyngeal and rectal swabs and blood samples were collected from 273 domestic cats during the second wave of COVID-19 infection in Latvia. Molecular prevalence was determined by using reverse transcriptase-polymerase chain reaction (RT-PCR). Serum samples were analysed via double antigen enzyme-linked immunosorbent assay targeting the antibody against the nucleocapsid protein of SARS-CoV-2. Positive swab samples were analysed using whole viral genome sequencing and subsequent phylogenetic analysis of the whole genome sequencing data of the samples was performed. RESULTS: The overall SARS-CoV-2 RT-PCR positivity and seroprevalence was 1.1% (3/273) and 2.6% (7/273), respectively. The SARS-CoV-2 genome sequences from three RT-PCR positive cats were assigned to the three common lineages (PANGOLIN lineage S.1.; B.1.177.60. and B.1.1.7.) circulating in Latvia during the particular period of time. CONCLUSIONS: These findings indicate that feline infection with SARS-CoV-2 occurred during the second wave of the COVID-19 pandemic in Latvia, yet the overall prevalence was low. In addition, it seems like no special 'cat' pre-adaptations were necessary for successful infection of cats by the common lineages of SARS-CoV-2.

Ryegrass mottle virus complete genome determination and development of infectious cDNA by combining two methods- 3' RACE and RNA-Seq.

Ryegrass mottle virus (RGMoV; genus: Sobemovirus) is a single-stranded positive RNA virus with a 30 nm viral particle size. It exhibits T = 3 symmetry with 180 coat protein (CP) subunits forming a viral structure. The RGMoV genome comprises five open reading frames that encode P1, Px, a membrane-anchored 3C-like serine protease, a viral genome-linked protein, P16, an RNA-dependent RNA polymerase, and CP. The RGMoV genome size varies, ranging from 4175 nt (MW411579.1) to 4253 nt (MW411579.1) in the deposited sequences. An earlier deposited RGMoV complete genome sequence of 4212 nt length (EF091714.1) was used to develop an infectious complementary DNA (icDNA) construct for in vitro gRNA transcription from the T7 promoter. However, viral infection was not induced when the transcribed gRNA was introduced into oat plants, indicating the potential absence of certain sequences in either the 5' or 3' untranslated regions (UTR) or both. The complete sequence of the 3' UTR was determined through 3' end RACE, while the 5' UTR was identified using high-throughput sequencing (HTS)-RNA-Seq to resolve the potential absences. Only the icDNA vector containing the newly identified UTR sequences proved infectious, resulting in typical viral infection symptoms and subsequent propagation of progeny viruses, exhibiting the ability to cause repeated infections in oat plants after at least one passage. The successful generation of icDNA highlighted the synergistic potential of utilizing both methods when a single approach failed. Furthermore, this study demonstrated the reliability of HTS as a method for determining the complete genome sequence of viral genomes.

Variations in the Relative Abundance of Gut Bacteria Correlate with Lipid Profiles in Healthy Adults.

The gut microbiome is a versatile system regulating numerous aspects of host metabolism. Among other traits, variations in the composition of gut microbial communities are related to blood lipid patterns and hyperlipidaemia, yet inconsistent association patterns exist. This study aims to assess the relationships between the composition of the gut microbiome and variations in lipid profiles among healthy adults. This study used data and samples from 23 adult participants of a previously conducted dietary intervention study. Circulating lipid measurements and whole-metagenome sequences of the gut microbiome were derived from 180 blood and faecal samples collected from eight visits distributed across an 11-week study. Lipid-related variables explained approximately 4.5% of the variation in gut microbiome compositions, with higher effects observed for total cholesterol and high-density lipoproteins. Species from the genera Odoribacter, Anaerostipes, and Parabacteroides correlated with increased serum lipid levels, whereas probiotic species like Akkermansia muciniphila were more abundant among participants with healthier blood lipid profiles. An inverse correlation with serum cholesterol was also observed for Massilistercora timonensis, a player in regulating lipid turnover. The observed correlation patterns add to the growing evidence supporting the role of the gut microbiome as an essential regulator of host lipid metabolism.

Genomic Characterization and Initial Insight into Mastitis-Associated SNP Profiles of Local Latvian Bos taurus Breeds.

Latvia has two local Bos taurus breeds-Latvian Brown (LBG) and Latvian Blue (LZG)-characterized by a good adaptation to the local climate, longevity, and high fat and protein contents in milk. Since these are desired traits in the dairy industry, this study investigated the genetic background of the LBG and LZG breeds and identified the genetic factors associated with mastitis. Blood and semen samples were acquired, and whole genome sequencing was then performed to acquire a genomic sequence with at least 35× or 10× coverage. The heterozygosity, nucleotide diversity, and LD analysis indicated that LBG and LZG cows have similar levels of genetic diversity compared to those of other breeds. An analysis of the population structure revealed that each breed clustered together, but the overall differentiation between the breeds was small. The highest genetic variance was observed in the LZG breed compared with the LBG breed. Our results show that SNP rs721295390 is associated with mastitis in the LBG breed, and SNPs rs383806754, chr29:43998719CG>C, and rs462030680 are associated with mastitis in the LZG breed. This study shows that local Latvian LBG and LZG breeds have a pronounced genetic differentiation, with each one suggesting its own mastitis-associated SNP profile.

Abundance and prevalence of ESBL coding genes in patients undergoing first line eradication therapy for Helicobacter pylori.

The spread of extended-spectrum beta-lactamases (ESBLs) in nosocomial and community-acquired enterobacteria is an important challenge for clinicians due to the limited therapeutic options for infections that are caused by these organisms. Here, we developed a panel of ESBL coding genes, evaluated the abundance and prevalence of ESBL encoding genes in patients undergoing H. pylori eradication therapy, and summarized the effects of eradication therapy on functional profiles of the gut microbiome. To assess the repertoire of known beta lactamase (BL) genes, they were divided into clusters according to their evolutionary relation. Primers were designed for amplification of cluster marker regions, and the efficiency of this amplification panel was assessed in 120 fecal samples acquired from 60 patients undergoing H. pylori eradication therapy. In addition, fecal samples from an additional 30 patients were used to validate the detection efficiency of the developed ESBL panel. The presence for majority of targeted clusters was confirmed by NGS of amplification products. Metagenomic sequencing revealed that the abundance of ESBL genes within the pool of microorganisms was very low. The global relative abundances of the ESBL-coding gene clusters did not differ significantly among treatment states. However, at the level of each cluster, classical ESBL producers such as Klebsiella sp. for blaOXY (p = 0.0076), Acinetobacter sp. for blaADC (p = 0.02297) and others, differed significantly with a tendency to decrease compared to the pre- and post-eradication states. Only 13 clusters were common across all three datasets, suggesting a patient-specific distribution profile of ESBL-coding genes. The number of AMR genes detected in the post-eradication state was higher than that in the pre-eradication state, which could be attributed, at least in part, to the therapy. This study demonstrated that the ESBL screening panel was effective in targeting ESBL-coding gene clusters from bacterial DNA and that minor differences exist in the abundance and prevalence of ESBL-coding gene levels before and after eradication therapy.

Wastewater-based prediction of COVID-19 cases using a random forest algorithm with strain prevalence data: A case study of five municipalities in Latvia.

Wastewater-based epidemiology (WBE) is a rapid and cost-effective method that can detect SARS-CoV-2 genomic components in wastewater and can provide an early warning for possible COVID-19 outbreaks up to one or two weeks in advance. However, the quantitative relationship between the intensity of the epidemic and the possible progression of the pandemic is still unclear, necessitating further research. This study investigates the use of WBE to rapidly monitor the SARS-CoV-2 virus from five municipal wastewater treatment plants in Latvia and forecast cumulative COVID-19 cases two weeks in advance. For this purpose, a real-time quantitative PCR approach was used to monitor the SARS-CoV-2 nucleocapsid 1 (N1), nucleocapsid 2 (N2), and E genes in municipal wastewater. The RNA signals in the wastewater were compared to the reported COVID-19 cases, and the strain prevalence data of the SARS-CoV-2 virus were identified by targeted sequencing of receptor binding domain (RBD) and furin cleavage site (FCS) regions employing next-generation sequencing technology. The model methodology for a linear model and a random forest was designed and carried out to ascertain the correlation between the cumulative cases, strain prevalence data, and RNA concentration in the wastewater to predict the COVID-19 outbreak and its scale. Additionally, the factors that impact the model prediction accuracy for COVID-19 were investigated and compared between linear and random forest models. The results of cross-validated model metrics showed that the random forest model is more effective in predicting the cumulative COVID-19 cases two weeks in advance when strain prevalence data are included. The results from this research help inform WBE and public health recommendations by providing valuable insights into the impact of environmental exposures on health outcomes.

Oral Microbiome Traits of Type 1 Diabetes and Phenylketonuria Patients in Latvia.

Some metabolic disorder treatments require patients to follow a specific diet or to consume supplements that, over time, can lead to oral microbiome alterations. Well-known disorders requiring such treatment are phenylketonuria (PKU), an inborn error of amino acid metabolism, and type 1 diabetes (T1D), a metabolic disorder that requires a specific diet regimen. Therefore, the aim of this study was to investigate the oral health and microbiome characteristics that might contribute to caries activity and periodontal disease risk in PKU and T1D patients. In this cross-sectional study, 45 PKU patients, 24 T1D patients, and 61 healthy individuals between the ages of 12 and 53 years were examined. Their anamnestic data and dental status were assessed by one dentist. Microbial communities were detected from saliva-isolated DNA using 16S rRNA gene V3-V4 sequencing on Illumina MiSeq sequencing platform. Results revealed that the PKU patient group displayed the highest number of extracted teeth (on average 1.34), carious teeth (on average 4.95), and carious activity (44.44% of individuals) compared to the T1D and CTRL groups. The lowest numbers of filled teeth (on average 5.33) and extracted teeth (on average 0.63) per individual were observed in T1D patients. Gingivitis appeared more often in the T1D group; however, possible risk of periodontal disease was seen in both the T1D and PKU patient groups. The highest number of differentially abundant genera was detected in the PKU group (n = 20), with enrichment of Actinomyces (padj = 4.17 × 10-22), Capnocytophaga (padj = 8.53 × 10-8), and Porphyromonas (padj = 1.18 × 10-5) compared to the CTRL group. In conclusion, the dental and periodontal health of PKU patients was found to be significantly inferior compared to T1D patients and healthy controls. T1D patients showed early signs of periodontal disease. Several genera that correlate with periodontal disease development were found in both groups, thus suggesting that T1D and PKU patients should seek early and regular dental advice and be educated about proper oral hygiene practices.

Evaluation of pathogenicity of Botrytis species isolated from different legumes.

Fungi of genus Botrytis are important pathogens of legumes, causing gray mold and chocolate spot diseases. The use of molecular methods to identify pathogens has resulted in the discovery of several new Botrytis species and new associations of pathogens with diseases. Thus, chocolate spot of faba bean is now associated with at least four species: B. fabae, B. cinerea, B. pseudocinerea and B. fabiopsis. Species of Botrytis differ in host plant, pathogenicity, fungicide resistance and other relevant properties that affect disease control. The aim of this study was to identify the species of Botrytis isolated from different legume crops and to evaluate their in vitro pathogenicity. Between 2014 and 2019, 278 isolates of Botrytis were obtained from infected legumes in Latvia. A phylogenetic analysis was carried out by sequencing three nuclear genes, RPB2, HSP60, and G3PDH, considered to be diagnostic for species in this genus. A set of 21 representative isolates was selected for pathogenicity tests on detached leaves of faba bean, field pea, lupin and soybean using 5-mm mycelium-agar plugs. The diameter of the formed lesions under the inoculated plug was measured crosswise each day. The datasets were subjected to analysis of variance with the split-plot design of the experiment and repeated-measures model. Six species were identified: B. cinerea, B. fabae, B. pseudocinerea, B. fabiopsis, B. euroamericana and B. medusae. In addition to the expected combinations of host and pathogen, naturally occurring infections of B. fabiopsis were found on chickpea, B. euroamericana on faba bean and B. medusae in lupin seeds. Species and isolate had significant effects on pathogenicity on all crops tested. Several isolates were pathogenic on two or more host species: two of B. pseudocinerea, two of B. cinerea, two of B. fabiopsis and the one of B. medusae. One isolate of B. pseudocinerea and two of B. fabiopsis caused primary lesions on all five host species. The results show that these Botrytis species have a broad host range that should be borne in mind when planning crop sequences and rotations.

Spatial variation of the gut microbiome in response to long-term metformin treatment in high-fat diet-induced type 2 diabetes mouse model of both sexes.

Antidiabetic drug metformin alters the gut microbiome composition in the context of type 2 diabetes and other diseases; however, its effects have been mainly studied using fecal samples, which offer limited information about the intestinal site-specific effects of this drug. Our study aimed to characterize the spatial variation of the gut microbiome in response to metformin treatment by using a high-fat diet-induced type 2 diabetes mouse model of both sexes. Four intestinal parts, each at the luminal and mucosal layer level, were analyzed in this study by performing 16S rRNA sequencing covering six variable regions (V1-V6) of the gene and thus allowing to obtain in-depth information about the microbiome composition. We identified significant differences in gut microbiome diversity in each of the intestinal parts regarding the alpha and beta diversities. Metformin treatment altered the abundance of different genera in all studied intestinal sites, with the most pronounced effect in the small intestine, where Lactococcus increased remarkably. The abundance of Lactobacillus was substantially lower in male mice compared to female mice in all locations, in addition to an enrichment of opportunistic pathogens. Diet type and intestinal layer had significant effects on microbiome composition at each of the sites studied. We observed a different effect of metformin treatment on the analyzed subsets, indicating the multiple dimensions of metformin's effect on the gut microbiome.

Epiphytic and Endophytic Fungi Colonizing Seeds of Two Poaceae Weed Species and Fusarium spp. Seed Degradation Potential In Vitro.

Fungi colonizing the surface and endosphere of two widespread Poaceae weed species, Avena fatua and Echinochloa crus-galli, were isolated to compare the taxonomic composition between the plant species, location, and year of the seed collection. The seed-degrading potential of Fusarium isolated from the seeds was tested by inoculating seeds of E. crus-galli with spore suspension. Molecular identification of epiphytic and endophytic fungal genera was performed by sequencing the ITS region of rDNA. Endophytes comprised of significantly lower fungal richness compared to epiphytes. A significant taxonomic overlap was observed between the endosphere and seed surface. The most abundant genera were Alternaria, Fusarium, Cladosporium, and Sarocladium. Analysis of similarities and hierarchical clustering showed that microbial communities were more dissimilar between the two plant species than between the years. Fusarium isolates with a high potential to infect and degrade E. crus-galli seeds in laboratory conditions belong to F. sporotrichioides and F. culmorum.

Batch-mode stimulation of hydrocarbons biodegradation in freshwater sediments from historically contaminated Aluksne lake.

Historical contamination of freshwater lakes with hydrocarbons (HC) due to anthropogenic activities represents a serious problem worldwide. This study was focused on hydrocarbons-contaminated sediments sampled in Lake Aluksne of glacial origin in Northeast Latvia. The batch experiments were aimed at evaluating the effect of bio-stimulation and bioaugmentation on the biodegradation of hydrocarbons in lake sediments (LS), as well as changes in microbial community structure and metabolic activity. The sediments were sampled from two points of the lake, 4-5 m and 8 m depth, respectively. These samples slightly differed by colour, count of diatoms, microbial respiration intensity and colour intensity of 2,6- dichlorophenolindophenol. Nevertheless, the trend in biodegradation activity was similar for both LS samples. The concentration of HC in LS during the 32-day incubation decreased in average from 465 mg/kg to 165 mg/kg and 117.5 mg/kg in the LS amended with nutrients and nutrients+microbial community, respectively. Different treatment types of LS resulted in differences in microbial respiration and HC-degrading activity. The Shotgun sequencing has revealed the main phyla present in the intact LS being Proteobacteria (48.8%), Actinobacteria (24.4%), Firmicutes (10.4%) and Bacteroidetes (5.0%). Incubation of LS for 32 days resulted in increasing abundance of Proteobacteria from 48.8% in the raw LS to 58-62%, mainly due to the increase of Betaproteobacteria. The functional annotation of gene families revealed that the most abundant gene families were associated with ATP binding, metal ion, magnesium ion, sulfur cluster, zinc ion binding, DNA binding and other essential components for cell functioning. The Shannon biodiversity index of culturable microorganisms in EcoPlates™ ranged from 2.28 to 2.85. The data obtained in this study indicated that the suggested approach is a potent remediation technology for further ex situ scaling up.

Understanding Bartonella-Associated Infective Endocarditis: Examining Heart Valve and Vegetation Appearance and the Role of Neutrophilic Leukocytes.

BACKGROUND: The endocardium and cardiac valves undergo severe impact during infective endocarditis (IE), and the formation of vegetation places IE patients at a heightened risk of embolic complications and mortality. The relevant literature indicates that 50% of IE cases exhibit structurally normal cardiac valves, with no preceding history of heart valve disease. Gram-positive cocci emerge as the predominant causative microorganisms in IE, while Gram-negative Bartonella spp., persisting in the endothelium, follow pathogenic pathways distinct from those of typical IE-causing agents. Employing clinical as well as advanced microbiological and molecular assays facilitated the identification of causative pathogens, and various morphological methods were applied to evaluate heart valve damage, shedding light on the role of neutrophilic leukocytes in host defense. In this research, the immunohistochemical analysis of neutrophilic leukocyte activation markers such as myeloperoxidase, neutrophil elastase, calprotectin, and histone H3, was performed. A distinct difference in the expression patterns of these markers was observed when comparing Bartonella spp.-caused and non-Bartonella spp.-caused IE. The markers exhibited significantly higher expression in non-Bartonella spp.-caused IE compared to Bartonella spp.-caused IE, and they were more prevalent in vegetation than in the valvular leaflets. Notably, the expression of these markers in all IE cases significantly differed from that in control samples. Furthermore, we advocated the use of 16S rRNA Next-Generation Sequencing on excised heart valves as an effective diagnostic tool for IE, particularly in cases where blood cultures yielded negative results. The compelling results achieved in this study regarding the enigmatic nature of Bartonella spp. IE's pathophysiology contribute significantly to our understanding of the peculiarities of inflammation and immune responses.

The Diversity of Fungi Involved in Damage to Japanese Quince.

In recent years, Japanese quince (Chaenomeles japonica) plantations in Latvia have increased. Interest in breeding Japanese quince is also known in other European countries and Russia. The occurrence and harmfulness of fungal diseases have become increasingly significant. However, there is a lack of overall information in the literature about the diversity of fungi afflicting C. japonica. In our study, we aimed to determine the diversity of fungi associated with C. japonica in Latvia, with the possibility of identifying the fungi that are most characteristically associated with certain parts of the plant. Our research was conducted from 2017 to 2019 in eight Japanese quince plantations in Latvia. Samples of plant parts with disease symptoms were collected. Pure cultures of fungi were obtained and identified using ITS region sequencing. We determined the relative density of identified genera of fungi, which were grouped using hierarchical cluster analysis depending on the plant part from which they were found. Various disease-like symptoms were observed and described. A total of 538 isolates of fungi were obtained that belong to 36 genera and represent different ecological niches. Fusarium, Alternaria, Botrytis, and Sarocladium were the genera most frequently found during our study. The number of identified cases of fungal genera differed depending on the part of the plant from which the fungi were obtained. However, it is not possible to relate a specific genus of fungus to only one certain part of a plant. Further research is needed to clarify the pathogenicity of detected fungi and the composition of species in the detected genera of fungi.

OverFlap PCR: A reliable approach for generating plasmid DNA libraries containing random sequences without a template bias.

Over the decades, practical biotechnology researchers have aimed to improve naturally occurring proteins and create novel ones. It is widely recognized that coupling protein sequence randomization with various effect screening methodologies is one of the most powerful techniques for quickly, efficiently, and purposefully acquiring these desired improvements. Over the years, considerable advancements have been made in this field. However, developing PCR-based or template-guided methodologies has been hampered by resultant template sequence biases. Here, we present a novel whole plasmid amplification-based approach, which we named OverFlap PCR, for randomizing virtually any region of plasmid DNA without introducing a template sequence bias.

Detection of SARS-CoV-2 RNA in wastewater and importance of population size assessment in smaller cities: An exploratory case study from two municipalities in Latvia.

Wastewater-based epidemiology (WBE) has regained global importance during the COVID-19 pandemic. The mobility of people and other factors, such as precipitation and irregular inflow of industrial wastewater, are complicating the estimation of the disease prevalence through WBE, which is crucial for proper crisis management. These estimations are particularly challenging in urban areas with moderate or low numbers of inhabitants in situations where movement restrictions are not adopted (as in the case of Latvia) because residents of smaller municipalities tend to be more mobile and less strict in following the rules and measures of disease containment. Thus, population movement can influence the outcome of WBE measurements significantly and may not reflect the actual epidemiological situation in the respective area. Here, we demonstrate that by combining the data of detected SARS-CoV-2 RNA copy number, 5-hydroxyindoleacetic acid (5-HIAA) analyses in wastewater and mobile call detail records it was possible to provide an accurate assessment of the COVID-19 epidemiological situation in towns that are small (COVID-19 28-day cumulative incidence r = 0.609 and 35-day cumulative incidence r = 0.89, p < 0.05) and medium-sized towns (COVID-19 21-day cumulative incidence r = 0.997, 28-day cumulative incidence r = 0.98 and 35-day cumulative incidence r = 0.997, p < 0.05). This is the first study demonstrating WBE for monitoring COVID-19 outbreaks in Latvia. We demonstrate that the application of population size estimation measurements such as total 5-HIAA and call detail record data improve the accuracy of the WBE approach.

Dominance of Fructose-Associated Fructobacillus in the Gut Microbiome of Bumblebees (Bombus terrestris) Inhabiting Natural Forest Meadows.

Bumblebees are key pollinators in agricultural landscapes. However, little is known about how gut microbial communities respond to anthropogenic changes. We used commercially produced colonies of buff-tailed bumblebees (Bombus terrestris) placed in three habitats. Whole guts (midgut, hindgut, and rectum) of B. terrestris specimens were dissected from the body and analyzed using 16S phylogenetic community analysis. We observed significantly different bacterial community composition between the agricultural landscapes (apple orchards and oilseed rape (Brassica napus) fields) and forest meadows, whereas differences in gut communities between the orchards and oilseed rape fields were nonsignificant. Bee-specific bacterial genera such as Lactobacillus, Snodgrassella, and Gilliamella dominated gut communities of B. terrestris specimens. In contrast, the guts of B. terrestris from forest meadows were dominated by fructose-associated Fructobacillus spp. Bacterial communities of workers were the most diverse. At the same time, those of males and young queens were less diverse, possibly reflecting greater exposure to the colony's inner environment compared to the environment outside the colony, as well as bumblebee age. Our results suggest that habitat quality, exposure to environmental microbes, nectar quality and accessibility, and land use significantly affect gut bacterial composition in B. terrestris.

First Report on the Latvian SARS-CoV-2 Isolate Genetic Diversity.

Remaining a major healthcare concern with nearly 29 million confirmed cases worldwide at the time of writing, novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has caused more than 920 thousand deaths since its outbreak in China, December 2019. First case of a person testing positive for SARS-CoV-2 infection within the territory of the Republic of Latvia was registered on 2nd of March 2020, 9 days prior to the pandemic declaration by WHO. Since then, more than 277,000 tests were carried out confirming a total of 1,464 cases of coronavirus disease 2019 (COVID-19) in the country as of 12th of September 2020. Rapidly reacting to the spread of the infection, an ongoing sequencing campaign was started mid-March in collaboration with the local testing laboratories, with an ultimate goal in sequencing as much local viral isolates as possible, resulting in first full-length SARS-CoV-2 isolate genome sequences from the Baltics region being made publicly available in early April. With 133 viral isolates representing ~9.1% of the total COVID-19 cases during the "first coronavirus wave" in the country (early March, 2020-mid-September, 2020) being completely sequenced as of today, here, we provide a first report on the genetic diversity of Latvian SARS-CoV-2 isolates.