16S Genes Research Articles

Gut microbiota transfer evidence from mother to newborn.

Early life microbiota is a risk factor for future diseases. The main purpose of this study was to investigate the transfer of gut microbiota from mother to newborn. A biological sample was collected from the anal mucosa of the pregnant women before delivery and from the newborns between 24 and 48h after delivery, as it was not possible to collect a meconium sample at that time. The microbiome of the samples was analyzed by sequencing the hypervariable regions V3-V4 of the 16S gene. To determine the likelihood of microbiota transfer from mother to newborn and examine the relationship with the mode of delivery, we utilized Fisher's exact test and odds ratio. A weighted transfer ratio was employed as a comprehensive measure of transfer. A total of 5767 ASVs were identified in newborn samples (n = 30) and 7253 in maternal samples (n = 30). In the analysis of transfer correlated with the mode of delivery, we observed significant ASVs (p < 0.05). Vaginal delivery showed a positive probability of transfer (OR = 2.184 and WTR = 1.852). We found a negative correlation (OR < 1) between the abundance of maternal ASVs and the likelihood of microbiota transfer to the newborn in both delivery modes. The relationship was inversely proportional for both cesarean section (log10 = - 0.2229) and vaginal delivery (log10 = - 0.1083), with statistical significance observed only for cesarean section (p = 0.0083). Conclusion:In our sample, the maternal gut microbiome was found to be associated with the infant gut microbiome, indicating evidence of ASV-specific transfer from the maternal microbiome to newborns. What is Known: • There is a relationship of early-life microbiota composition with future health outcomes. What is New: • This was the first study to evaluate maternal gut microbiota transfer to newborns in Brazil.

Molecular surveillance reveals a potential hotspot of tick-borne disease in Yakeshi City, Inner Mongolia

A molecular surveillance of tick-borne diseases was performed in Hulunbuir City, Inner Mongolia. A total of 149 ticks including three species (Ixodes persulcatus, Haemaphysalis concinna, and Dermacentor silvarum) were collected. As many as 11 tick-borne bacterial pathogens were identified in them. Some of them have high positive rates. For example, Candidatus Rickettsia tarasevichiae was detected with a high prevalence of 72.48%, while Candidatus Lariskella sp. was detected in 31.54% of ticks. For both Rickettsia raoultii and Anaplasma phagocytophilum, two distinct genotypes were identified based on their phylogenetic trees based on 16S rRNA, gltA, and groEL sequences. Remarkable genetic diversity was also observed for 16S and flaB genes of Borreliella garinii, an agent of Lyme disease. Rickettsia heilongjiangensis causing Far-Eastern spotted fever (2.68%, 4/149), Ehrlichia muris causing human ehrlichiosis (4.70%, 7/149), Borrelia miyamotoi causing relapsing fever (2.01%, 3/149), and Borreliella afzelii causing Lyme disease (2.01%, 3/149) were also detected. Additionally, a previously uncharacterized Anaplasma species closely related to Anaplasma ovis was identified. Herein we name it “Candidatus Anaplasma mongolica”. Based on these results, we propose that Yakeshi City might be a potential hotspot of tick-borne diseases.

Another new species (and its not over yet) of Phyllodytes Wagler, 1930 (Anura, Hylidae) from the Atlantic Forest of southern Bahia, northeastern Brazil.

The genus Phyllodytes, endemic to the Atlantic Forest, stands out for its life cycle being closely associated with bromeliads. Since the 2000s, the number of species in the group has more than doubled and the number is still increasing, a fact proven here with the description of Phyllodytes iuna sp. nov. This species, herein described using morphological and molecular evidence, is endemic to the Atlantic Forest of southern Bahia, in the northern portion of the Atlantic Forest Central Corridor. Phyllodytes iuna differs from both its sister species and most of its congeners by the color pattern on the dorsum and by the presence of a single tubercle in the tibiotarsal region. The genetic distance for a fragment of the 16S gene ranged from 4.4% to 5.0% between Phyllodytes iuna and its sister species (P. brevirostris and P. edelmoi), and from 5.8% to 14.2% with its congeners. The description of a new species of Phyllodytes serves as a reminder that our understanding of the groups diversity is far from complete. Despite the alarming rates of deforestation, the Atlantic Forest continues to hold significant untapped potential for unexplored biodiversity.

Discovery, Identification, and Insecticidal Activity of an Aspergillus flavus Strain Isolated from a Saline-Alkali Soil Sample.

Aphids are one of the most destructive pests in agricultural production. In addition, aphids are able to easily develop resistance to chemical insecticides due to their rapid reproduction and short generation periods. To explore an effective and environmentally friendly aphid control strategy, we isolated and examined a fungus with aphid-parasitizing activity. The strain (YJNfs21.11) was identified as Aspergillus flavus by ITS, 28S, and BenA gene sequence analysis. Scanning electron microscopy and transmission electron microscopy revealed that the infection hyphae of 'YJNfs21.11' colonized and penetrated the aphid epidermal layer and subsequently colonized the body cavity. Field experiments showed that 'YJNfs21.11' and its fermentation products exerted considerable control on aphids, with a corrected efficacy of 96.87%. The lipase, protease, and chitinase secreted by fungi help aphid cuticle degradation, thus assisting spores in completing the infection process. Additionally, changes were observed in the mobility and physical signs of aphids, with death occurring within 60 h of infection. Our results demonstrate that A. flavus 'YJNfs21.11' exhibits considerable control on Aphis gossypii Glover and Hyalopterus arundimis Fabricius, making it a suitable biological control agent.

First genetic insights of Gonatodescaudiscutatus (Reptilia, Gekkota) in the Galapagos Islands and mainland Ecuador.

Studies on genetic variability amongst native and introduced species contribute to a better understanding of the genetic diversity of species along their autochthonous distribution and identify possible routes of introduction. Gonatodescaudiscutatus is a gecko native to western Ecuador and introduced to the Galapagos Islands. Despite being a successful species in human-modified habitats along its native and non-native ranges, neither the colonisation process nor the genetic diversity of this gecko is known. In this study, we analysed 55 individuals from 14 localities in western Ecuador and six localities in San Cristobal Island, Galapagos - the only island with a large, self-sustaining population. We amplified and analysed the genetic variability of two nuclear genes (Cmos and Rag2) and one mitochondrial gene (16S). Cmos and Rag2 sequences presented little to none genetic variability, while 16S allowed us to build a haplotype network. We identified nine haplotypes across mainland Ecuador, two of which are also present in Galapagos. Low genetic diversity between insular and continental populations suggests that the introduction of G.caudiscutatus on the Islands is relatively recent. Due to the widespread geographical distribution of mainland haplotypes, it was not possible to determine the source population of the introduction. This study represents the first exploration of the genetic diversity of Gonatodescaudiscutatus, utilising genetic tools to gain insights into its invasion history in the Galapagos.

Expanded Multiplexing on Sensor-Constrained Microfluidic Partitioning Systems.

Microfluidics can split samples into thousands or millions of partitions, such as droplets or nanowells. Partitions capture analytes according to a Poisson distribution, and in diagnostics, the analyte concentration is commonly inferred with a closed-form solution via maximum likelihood estimation (MLE). Here, we present a new scalable approach to multiplexing analytes. We generalize MLE with microfluidic partitioning and extend our previously developed Sparse Poisson Recovery (SPoRe) inference algorithm. We also present the first in vitro demonstration of SPoRe with droplet digital PCR (ddPCR) toward infection diagnostics. Digital PCR is intrinsically highly sensitive, and SPoRe helps expand its multiplexing capacity by circumventing its channel limitations. We broadly amplify bacteria with 16S ddPCR and assign barcodes to nine pathogen genera by using five nonspecific probes. Given our two-channel ddPCR system, we measured two probes at a time in multiple groups of droplets. Although individual droplets are ambiguous in their bacterial contents, we recover the concentrations of bacteria in the sample from the pooled data. We achieve stable quantification down to approximately 200 total copies of the 16S gene per sample, enabling a suite of clinical applications given a robust upstream microbial DNA extraction procedure. We develop a new theory that generalizes the application of this framework to many realistic sensing modalities, and we prove scaling rules for system design to achieve further expanded multiplexing. The core principles demonstrated here could impact many biosensing applications with microfluidic partitioning.

Musicians in the marsh: a new species of music frog (Anura: Ranidae: Nidirana) from Arunachal Pradesh, India.

We describe a new species of ranid frog of the genus Nidirana from northeast India based on morphological, molecular and acoustic evidence. The new species is phenotypically distinct from its congeners by a combination of morphological characters: body robust with SVL 46.559.1 mm (n= 3) in adult males and SVL 60.666.0 mm (n= 2) in adult females; a pair of subgular vocal sacs and two patches of nuptial pad on the first finger in adult males; toe tips slightly dilated and oval; circum-marginal grooves present on all toes; dorsal skin with scattered small tubercles. A pale cream-coloured mid-dorsal line from the snout tip to the vent is present. Phylogenetically, the new species differs from its congeners by a genetic divergence of 3.48.0% and 7.712.4% in 16S and COI genes respectively. Furthermore, the new species can be differentiated from its congeners by its advertisement call, which consists of two different types of notes, call duration (0.580.92 s) and dominant frequency of the call (473.7 Hz). The discovery of a new species validates the presence of the genus Nidirana from India and emphasizes the importance of exploring specialized habitats such as marshlands, which are often overlooked.

First Report of Powdery Mildew Caused by Podosphaera xanthii on Vernonia cinerea in China.

Vernonia cinerea (L.)Less. is an annual herbaceous plant of the Asteraceae family, which is widely distributed throughout Southeast Asia, India, and the tropical and subtropical regions of China. This herb is known to contain various bioactive compounds and is commonly used in traditional system of medicine in China and India (Singh et al. 2021). During the spring of 2022 and 2023, powdery mildew symptoms were observed on 70% of V. cinerea plants on the Hainan Medical University campus (19° 58' 53″ N; 110° 19' 47″ E) in Haikou, Hainan Province, China. Powdery mildew colonies covered the leaf surfaces and stems of affected plants, causing discoloration and defoliation. Mycelia were superficial and hyphal appressoria were nipple-shaped. Conidiophores (n =30) were unbranched, cylindrical, 104 to 188 × 11 to 15 µm, and produced three to five immature conidia in chains with a crenate outline. Foot cells (n =30) were cylindrical, straight or sometimes curved at the base, and 32 to 57 µm long. Conidia (n =100) were ellipsoid-ovoid to doliiform, 19 to 45 ×16 to 26 m (length/width ratio = 1.2 to 2.4), with well-developed fibrosin bodies, and produced germ tubes from the lateral position. Based on these morphological characteristics, the pathogen was provisionally identified as Podosphaera xanthii (Braun and Cook 2012). The teleomorph was not observed. A specimen was deposited in the Hainan Medical University Plant Pathology Herbarium as HMVC-22. To confirm the genus identification and ascertain a putative species, genomic DNA was extracted from mycelium, conidiophores, and conidia using a fungal DNA kit (Omega Bio-Tek, USA). The rDNA internal transcribed spacer (ITS) region was amplified with primers ITS1/ITS4 (White et al. 1990) and sequenced directly. The resulting 577-bp sequence was deposited in GenBank (accession no. OP765400). A BLASTn search in GenBank of this sequence showed 100% similarity with the ITS sequences of P. xanthii isolates from China (MT260063, MT739423 and MT131253), Thailand (LC270780), and Vietnam (KM260731). Additionally, the 28S rDNA region was amplified using the primer pairs NL1 and NL4 (O´Donnell 1993; accession no. OP765401). This region shared 100% similarity with P. xanthii isolates (LC371333, LC270780, and AB936277) as well. Powdery mildew from Hainan sample belonged to the P. xanthii group with strong bootstrap values support 99% in maximum likelihood phylogenetic tree based on ITS and 28S gene sequences. To confirm pathogenicity, five healthy potted plants of V. cinerea were inoculated by gently pressing a powdery mildew-infected leaf onto 15 young leaves. Five non-inoculated plants served as controls. All plants were maintained in a greenhouse at 24 to 30°C, 70% relative humidity, with a 16-h photoperiod. After 7 days, inoculated leaves showed powdery mildew symptoms whereas no symptoms were observed on control plants. The fungal colonies observed on inoculated plants were morphologically identical to those found on the originally infected leaves collected from Hainan Province. Based on the morphological characteristics and molecular identification, the fungus was identified as P. xanthii. In different countries and regions, Erysiphe cichoracearum has been previously reported on some Vernonia species, including V. Galamensis from southern parts of Ethiopia (Hundesa and Mekonnen 2017). To our knowledge, this is the first record of P. xanthii infecting V. cinerea in China. We are concerned that the pathogen could become a threat to the widespread planting of V. cinerea in the future.

In silico prospecting of the mtDNA of Macrobrachium amazonicum from transcriptome data

BackgroundMacrobrachium amazonicum is a freshwater prawn widely distributed in South America that is undergoing speciation, so the denomination “M. amazonicum complex” is used for it. The mitochondrial cytochrome c oxidase subunit I (COI) gene has been used to elucidate this speciation, but heteroplasmies and pseudogenes have been recorded, making separation difficult. Obtaining genes from cDNA (RNA) rather than genomic DNA is an effective tool to mitigate those two types of occurrences. The aim of this study was to assemble in silico the mitochondrial DNA (mtDNA) of the Amazonian coastal population of M. amazonicum inhabiting the state of Pará.ResultsSequences were obtained from the prawn’s transcriptome using the de novo approach. Six libraries of cDNA from the androgen gland, hepatopancreas, and muscle tissue were used. The mtDNA of M. amazonicum was 14,960 bp in length. It contained 13 protein-coding genes, 21 complete transfer RNAs, and the 12S and 16S subunits of ribosomal RNA. All regions were found on the light strand except tRNAGln, which was on the heavy strand. The control region (D-loop) was not recovered, making for a gap of 793 bp. The cladogram showed the formation of the well-defined Macrobrachium clade, with high support value in the established branches (91–100). The three-dimensional spatial conformation of the mtDNA-encoded proteins showed that most of them were mainly composed of major α-helices that typically shows in those proteins inserted in the membrane (mitochondrial).ConclusionsIt was possible to assemble a large part of the mitochondrial genome of M. amazonicum in silico using data from other genomes deposited in GenBank and to validate it through the similarities between its COI and 16S genes and those from animals of the same region deposited in GenBank. Depositing the M. amazonicum mtDNA sequences in GenBank may help solve the taxonomic problems recorded for the species, in addition to providing complete sequences of candidate coding genes for use as biomarkers in ecological studies.

Rare and abundant bacterial communities in poplar rhizosphere soils respond differently to genetic effects

Interactions between plants and soil microbes are important to plant hybrid breeding under global change. However, the relationship between host plants and rhizosphere soil microorganisms has not been fully elucidated. Understanding the rhizosphere microbial structure of parents and progenies would provide a deeper insight into how genetic effects modulate the relationship between plants and soil. In this study, two family groups of poplar trees (A: parents and their two progenies; B: parents and their one progeny) with different genetic backgrounds (including seven genotypes) were selected from a common garden, and their rhizobacterial communities were analyzed to explore parent-progeny relationships. Our results showed significant differences in phylogenetic diversity, the number of 16S genes and the structure of rhizosphere bacterial communities (Adonis: R2 = 0.166, P < 0.01) between different family groups. Rhizosphere bacterial community structure was significantly dominated by genetic effects. Compared with abundant taxa, genetic effects were more powerful drivers of rare taxa. In addition, bacterial communities of hybrid progenies were all significantly more similar to their parents compared to the other group of parents, especially among rare taxa. The two poplar family groups exhibited differences between their rhizosphere bacterial co-occurrence networks. Group B had a relatively complex network with 2380 edges and 468 nodes, while group A had 1829 edges and 304 nodes. Soil organic carbon and carbon to nitrogen ratio (C/N) also influenced the rhizosphere bacterial community assembly. This was especially true for soil C/N, which explained 23 % of the β-nearest taxon index (βNTI) variation in rare taxa. Our results reveal the relationship of rhizosphere microorganisms between parents and progenies. This can help facilitate an understanding of the combination of plant breeding with microbes resource utilization and provide a theoretical basis for scientific advancement to support the development of forestry industry.

Enhanced immunogenicity elicited by a novel DNA vaccine encoding the SARS-CoV-2 S1 protein fused to the optimized flagellin of Salmonella typhimurium in mice.

The development of safe and efficient vaccines against coronavirus disease 2019 (COVID-19) is necessary for global public health. The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major target for the COVID-19 vaccines. Among the available vaccine types, DNA vaccines are a promising alternative to conventional vaccines. Here, we developed a novel DNA vaccine containing the SARS-CoV-2 S1 subunit and an optimized flagellin adjuvant (FliCΔD2D3) from Salmonella typhimurium. The S1 gene and S1-fliCΔD2D3 fusion gene were codon optimized and then cloned into the antibiotic-resistance-gene-free vector asd-pVAX1, producing asd-pVAX1-S1 and asd-pVAX1-S1-fliCΔD2D3, respectively. The expression of S1 and S1-FliCΔD2D3 proteins was confirmed after transient expression in HEK293T cells. After in vitro characterization of the vaccine candidates, we evaluated the humoral and cellular immune responses induced in BALB/c mice. The DNA vaccine induced a long-lasting humoral response for approximately 4 months, and the elicited serum antibodies efficiently blocked the binding of the SARS-CoV-2 S1 receptor-binding domain to its entry receptor (angiotensin-converting enzyme 2). Moreover, the DNA vaccine induced strong Th1/Th2-mixed immune responses, as demonstrated by the similar levels of S1-specific immunoglobulin (Ig) G1 and IgG2a in the serum and the significantly elevated Th1-type (IFN-γ and TNF-α) and Th2-type (IL-4 and IL-6) cytokine expression from splenic lymphocytes. Importantly, asd-pVAX1-S1-fliCΔD2D3 immunization in mice could significantly enhance the levels of immune responses compared to asd-pVAX1-S1. This study provides crucial information regarding the selection of a safer DNA vector and a new adjuvant for the vaccine development of SARS-CoV-2 and other infectious diseases. IMPORTANCE The development of safe and effective vaccines is needed to control the transmission of coronavirus disease 2019 (COVID-19). Synthetic DNA vaccines represent a promising platform in response to such outbreaks. Here, DNA vaccine candidates were developed using an optimized antibiotic-resistance gene-free asd-pVAX1 vector. An optimized flagellin (FliC) adjuvant was designed by fusion expression to increase the immunogenicity of the S1 antigen. S1 and S1-FliCΔD2D3 proteins were strongly expressed in mammalian cells. The FliCΔD2D3-adjuvanted DNA vaccine induced Th1/Th2-mixed immune responses and high titers of neutralizing antibodies. This study provides crucial information regarding the selection of a safer DNA vector and adjuvant for vaccine development. Our FliCΔD2D3-adjuvanted S1 DNA vaccine is more potent at inducing both humoral and cellular immune responses than S1 alone. This finding provides a new idea for the development of novel DNA vaccines against COVID-19 and could be further applied for the development of other vaccines.

Multiple evolutionary transitions of reproductive strategies in a phylum of aquatic colonial invertebrates.

Parental care is considered crucial for the enhanced survival of offspring and evolutionary success of many metazoan groups. Most bryozoans incubate their young in brood chambers or intracoelomically. Based on the drastic morphological differences in incubation chambers across members of the order Cheilostomatida (class Gymnolaemata), multiple origins of incubation were predicted in this group. This hypothesis was tested by constructing a molecular phylogeny based on mitogenome data and nuclear rRNA genes 18S and 28S with the most complete sampling of taxa with various incubation devices to date. Ancestral character estimation suggested that distinct types of brood chambers evolved at least 10 times in Cheilostomatida. In Eucratea loricata and Aetea spp. brooding evolved unambiguously from a zygote-spawning ancestral state, as it probably did in Tendra zostericola, Neocheilostomata, and 'Carbasea' indivisa. In two further instances, brooders with different incubation chamber types, skeletal and non-skeletal, formed clades (Scruparia spp., Leiosalpinx australis) and (Catenicula corbulifera (Steginoporella spp. (Labioporella spp., Thalamoporella californica))), each also probably evolved from a zygote-spawning ancestral state. The modular nature of bryozoans probably contributed to the evolution of such a diverse array of embryonic incubation chambers, which included complex constructions made of polymorphic heterozooids, and maternal zooidal invaginations and outgrowths.

Isolation and Identification of Green Microalgae from Northern Jordan

In light of the rapid and extreme changes in climate and the steady depletion of natural resources, there is an urgent need to find innovative and sustainable solutions to these problems. Microorganisms such as microalgae can offer viable solutions to these challenges. Proper investment in such organisms requires the identification of the algal species that inhabit the region. Therefore, this study aimed to isolate and molecularly characterize green microalgae that inhabit freshwater at different locations in the governorates of Irbid and Ajloun in the northern region of Jordan. Water samples collected from these regions were used to isolate single colonies, some of which exhibited different morphological characteristics. Genomic DNA was extracted from the isolates and used as a template for PCR amplification of the 18S ribosomal DNA gene (18S rDNA) and the internal transcribed spacer (ITS) region. Phylogenetic trees were constructed based on 18S rDNA and ITS PCR product sequences, which were used to identify the isolates at the genus level. The obtained isolates belonged to three genera; Coelastrella, Desmodesmus, and Monoraphidium. The latter species has not been previously reported in Jordan.

Experimental erosion of microbial diversity decreases soil CH4 consumption rates.

Biodiversity-ecosystem functioning (BEF) experiments have predominantly focused on communities of higher organisms, in particular plants, with comparably little known to date about the relevance of biodiversity for microbially driven biogeochemical processes. Methanotrophic bacteria play a key role in Earth's methane (CH4 ) cycle by removing atmospheric CH4 and reducing emissions from methanogenesis in wetlands and landfills. Here, we used a dilution-to-extinction approach to simulate diversity loss in a methanotrophic landfill cover soil community. Replicate samples were diluted 101 -107 -fold, preincubated under a high CH4 atmosphere for microbial communities to recover to comparable size, and then incubated for 86 days at constant or diurnally cycling temperature. We hypothesize that (1) CH4 consumption decreases as methanotrophic diversity is lost, and (2) this effect is more pronounced under variable temperatures. Net CH4 consumption was determined by gas chromatography. Microbial community composition was determined by DNA extraction and sequencing of amplicons specific to methanotrophs and bacteria (pmoA and 16S gene fragments). The richness of operational taxonomic units (OTU) of methanotrophic and nonmethanotrophic bacteria decreased approximately linearly with log-dilution. CH4 consumption decreased with the number of OTUs lost, independent of community size. These effects were independent of temperature cycling. The diversity effects we found occured in relatively diverse communities, challenging the notion of high functional redundancy mediating high resistance to diversity erosion in natural microbial systems. The effects also resemble the ones for higher organisms, suggesting that BEF relationships are universal across taxa and spatial scales.

Environmental DNA metabarcoding of water samples as a tool for monitoring Iberian freshwater fish composition.

Environmental DNA (eDNA) metabarcoding has been increasingly used to monitor the community assemblages of a wide variety of organisms. Here, we test the efficacy of eDNA metabarcoding to assess the composition of Iberian freshwater fishes, one of the most endangered groups of vertebrates in Spain. For this purpose, we sampled 12 sampling sites throughout one of Spain's largest basins, the Duero, which is home to approximately 70% of the genera and 30% of the primary freshwater fish in Spain. We sampled these sampling sites in the summer by using electrofishing, a traditional sampling method, and eDNA metabarcoding of river water samples using the mitochondrial 12S rRNA gene (12S) as a marker. We also resampled four of these sampling sites in autumn by eDNA. We compared the results obtained through eDNA metabarcoding with those of electrofishing surveys (ones conducted for the present study and past ones) and assessed the suitability of 12S as an eDNA metabarcoding marker for this group of freshwater fishes. We found that the 12S fragment, analysed for 25 Iberian species, showed sufficient taxonomic resolution to be useful for eDNA approaches, and even showed population-level differences in the studied populations across the tissue samples for Achondrostoma arcasii. In most cases, a greater number of species was detected through eDNA metabarcoding than through electrofishing. Based on our results, eDNA metabarcoding is a powerful tool to study the freshwater fish composition in the Iberian Peninsula and to unmask cryptic diversity. However, we highlight the need to generate a local genetic database for 12S gene for such studies and to interpret the results with caution when studying only mitochondrial DNA. Finally, our survey shows that the high detection sensitivity of eDNA metabarcoding and the non-invasiveness of this method allows it to act as a detection system for species of low abundance, such as early invasive species or species in population decline, two key aspects of conservation management of Spanish freshwater fishes.

A large scale temporal and spatial environmental DNA biodiversity survey of marine vertebrates in Brazil following the Fundão tailings dam failure

Seawater contains a wealth of genetic information, representing the biodiversity of numerous species residing within a particular marine habitat. Environmental DNA (eDNA) metabarcoding offers a cost effective, non-destructive method for large scale monitoring of environments, as diverse taxonomic groups are detected using metabarcoding assays. A large-scale eDNA monitoring program of marine vertebrates was conducted across three sampling seasons (Spring 2018, Autumn 2019; Spring 2019) in coastal waters of Brazil. The program was designed to investigate eDNA as a testing method for long term monitoring of marine vertebrates following the Fundão tailings dam failure in November 2015. While no baseline samples were available prior to the dam failure there is still value in profiling the taxa that use the impacted area and the trajectory of recovery. A total of 40 sites were sampled around the mouths of eight river systems, covering approximately 500 km of coastline. Metabarcoding assays targeting the mitochondrial genes 16S rRNA and COI were used to detect fish, marine mammals and elasmobranchs. We detected temporal differences between seasons and spatial differences between rivers/estuaries sampled. Overall, the largest eDNA survey in Brazil to date revealed 69 families from Class Actinopterygii (fish), 15 species from Class Chondrichthyes (sharks and rays), 4 species of marine and estuarine mammals and 23 species of conservation significance including 2 species of endangered dolphin. Our large-scale study reinforces the value eDNA metabarcoding can bring when monitoring the biodiversity of coastal environments and demonstrates the importance of collection of time-stamped environmental samples to better understand the impacts of anthropogenic activities.

The pH tolerance range of the airborne species Tetracystis vinatzeri (Chlorophyceae, Chlamydomonadales)

ABSTRACT To decipher the dispersal range of the cosmopolitan green microalga Tetracystis vinatzeri, two airborne-collected strains were barcoded using the 18S ribosomal DNA gene (18S) and the mitochondrial cytochrome c oxidase I subunit (COI), and their survival capacities were investigated across a stepwise acidic to alkaline pH gradient. Genetic fingerprinting confirmed the identity of the strains as representing the same biological species. Investigations of the tolerance of T. vinatzeri to pH levels refined the delimitation of its ecological niche. T. vinatzeri showed optimal growth in freshwater habitats at pH 4.3–9.6 and strains could cope with exposure to slightly more acidic (down to 3.8) and alkaline (up to 10.4) conditions. However, organismal survival was drastically affected by exposure to pH < 3 and > 11. Morphological observations showed that T. vinatzeri could better cope with exposure to acidic pH, exhibiting less cell membrane damage. Genetic analysis showed that the two airborne strains shared the same haplotype, suggesting that they either originated from the same air mass or that the haplotype has a widespread distribution. Further phylogeographic and physiological investigations are required to define the spreading dynamic of T. vinatzeri.

A new species of Xenophrys (Amphibia, Anura, Megophryidae) from southern Tibet, China.

A new species of Xenophrys is described from Yadong County, Tibet Autonomous Region, China based on morphological and molecular evidence. Phylogenetic analyses based on the mitochondrial genes 16S rRNA and COI indicated that this new species represents an independent lineage and the minimum p-distance based on 16S rRNA between this species and its congeners is 4.4%. Additionally, the new species is distinguished from its congeners by a combination of the following morphological characters: (1) small body size, SVL 17.9-22.2 mm in adult males and SVL 23.4 mm in the single adult female; (2) tympanum indistinct, supratympanic fold distinct; (3) canthus rostralis well-developed, snout tip far beyond the margin of the lower lip; (5) pupil vertical; (6) vomerine teeth present, maxillary teeth present; (7) tongue notched posteriorly; (8) supernumerary tubercles absent, subarticular, metacarpal and metatarsal tubercles indistinct; (9) relative finger lengths I < II < IV < III, finger tips rounded, slightly expanded relative to digit widths; (10) toes with narrow lateral fringes and tarsal folds; (11) a dark triangular marking with light edge between eyes, a dark ")("-shaped marking, with light edge, present on center of dorsum, pectoral glands on sides of the breast.

GADPO: Genetic Algorithm based on Dominance for Primer Optimization

The amplification of the 16S ribosomal RNA gene through the polymerase chain reaction (PCR) is the main approach to profile bacterial communities. This gene is a widely used marker with a composition that allows the identification of microorganisms at the genus or species levels. The correct performance of a PCR assay depends on the properties of the chosen set of primers that match with the target 16S sequences, allowing the amplification by a DNA polymerase. For this reason, optimizing the design of primers attending to such multiple properties is crucial to ensure the specificity and robustness of this process. However, only one multiobjective proposal exists that addresses the optimization of primer design targeted to the 16S gene amplification. Herein, we propose a novel approach for multiobjective primer optimization based on a mutation-based genetic algorithm with three new problem-aware mutation operators, each aimed at covering one of the three objectives that compose the problem (efficiency, coverage, and variability). The proposed algorithm has been tested on 5 real datasets of bacterial 16S gene sequences. The results have been evaluated using 4 quality metrics, showing that our approach achieves statistically significant improvements with regard to the reference multiobjective approach in the field.

Helminths of small rodents (Heteromyidae and Cricetidae) in the Yucatan Peninsula, Mexico: an integrative taxonomic approach to their inventory.

In this survey, we inventoried the helminths of heteromyid and cricetid rodents captured in the Yucatan Peninsula from 2017 to 2019. Helminths were identified using morphological techniques (clearing, staining, and scanning electron microscopy). Also, the 28S rRNA gene of individuals from several helminth taxa was successfully amplified and sequenced. To confirm the identification at the generic level, and in some cases at the specific level, and the genealogical relationships of the parasites, phylogenetic analyses were performed with the new 28S sequences. We identified 22 species of helminths including three trematodes (Brachylaimidae, Dicrocoeliidae, and Microphallidae), five cestodes (Davaineidae, Hymenolepididae, and Taeniidae), and 14 nematodes (Trichuridae, Ancylostomatidae, Ornithostrongylidae, Heligmonellidae, and Oxyuridae) from Heteromys gaumeri (Heteromyidae), Ototylomys phyllotis, Oligoryzomys fulvescens, Peromyscus yucatanicus, Sigmodon toltecus, and Reithrodontomys gracilis (Cricetidae). The overall frequency of infection in small rodents was 84.1% (143/170); all specimens of H. gaumeri, S. toltecus and Ol. fulvescens were infected with helminths. In total, we provided 46 new sequences of the 28S gene from 17 species of helminths. Seven species are likely undescribed species, six are reported for the first time in rodents from Mexico, and 12 are new host records in the Americas. Before this study, 87 taxa of helminths had been reported from 35 cricetid and 12 heteromyid species in 21 Mexican states. Our findings increase to 93 the helminth taxa in these rodents, and to 36 the cricetid species parasitized by helminths. This large scale-survey is the first to use an integrative approach to inventory the helminths of wild small rodents in Mexico.