Partial Sequence Research Articles

Resistance to acetyl-CoA carboxylase (ACCase)-inhibiting herbicides in Lolium multiflorum Lam. populations of Argentina.

Italian ryegrass (Lolium multiflorum Lam.) is one of the most troublesome grass weeds in Argentina. The extensive and repetitive use of acetyl-CoA carboxylase (ACCase)-inhibiting herbicides has induced resistance in this weed species. The objectives of this study were to quantify the resistance levels to ACCase-inhibiting herbicides in two resistant populations and to identify the target-site mutations associated with their resistance. Two resistant Italian ryegrass populations, Roldán and H2, were studied. Roldán was a suspected haloxyfop-resistant population, located in a wheat field from Santa Fe province with a history of ACCase-inhibiting herbicide use. The H2 population was obtained from the susceptible Hernandarias population (H0) after two cycles of selection with the herbicide quizalofop-ethyl. Whole-plant dose-response assays revealed that the resistant populations exhibited a high resistance to haloxyfop, with resistance factors (RF) exceeding 97-fold. Additionally, both populations showed a moderate resistance to pinoxaden (RF > 7), while maintaining susceptibility to clethodim. Partial chloroplastic ACCase sequences revealed isoleucine-to-asparagine substitution at position 2041 (Ile-2041-Asn) in both resistant populations. This work provides a better understanding of cross-resistance to ACCase-inhibiting herbicides in L. multiflorum populations and represents the first report of the target-site mutation Ile-2041-Asn conferring resistance in populations from Argentina. © 2024 Society of Chemical Industry.

Reaching new lands: Updating the distribution of Angiostrongylus cantonensis in South America with the first record in Argentina.

Angiostrongylus cantonensis, commonly known as the rat lungworm, is a metastrongyloid nematode found primarily not only in tropical and subtropical regions but also in temperate areas and considered the leading cause of eosinophilic meningitis in humans. Synanthropic rodents such as Rattus norvegicus and Rattus rattus are the most frequent definitive hosts of this parasite. The presence of this parasite was detected in the pulmonary arteries of three specimens of R. norvegicus in the city of Buenos Aires representing the species' southernmost known record in natural hosts. Species confirmation was achieved through partial sequences of 18S and COI genes. By comparing the COI gene sequences with those available in GenBank through the construction of a haplotype network, we obtained that the analysed specimen presents high similarity with those reported in Japan and Southeast Asia. All infected rats were captured in an area surrounding a port with significant import and export activity, suggesting that A. cantonensis may have been introduced through commercial ships. Specifically, the parasite was detected in a neighbourhood with vulnerable socio-economic conditions and in a nature reserve, which exhibit biotic and abiotic characteristics conducive to sustaining high-density rat populations, scattered waste, areas of spontaneous vegetation, debris accumulation and flooded areas or lagoons offering suitable habitats for intermediate hosts such as snails. Thus, the close proximity of the port to these sites creates a favourable ecological context for the establishment of A. cantonensis. This study shows the need to conduct research to detect A. cantonensis in non-endemic areas but with the characteristics that promote its arrival and development of its life cycle in order to implement control measures to prevent expansion of this parasite and its transmission to humans and other animals.

Assessing the Potential of Tortistilus (Hemiptera: Membracidae) from Northern California Vineyards as Vector Candidates of Grapevine Red Blotch Virus

Ceresini treehoppers are present in northern California vineyard ecosystems, including the closely related Spissistilus and Tortistilus (Hemiptera: Membracidae). These membracids are not direct pests of wine grapes, but S. festinus is a vector of grapevine red blotch virus (GRBV). No information is available on the ability of Tortistilus spp. to transmit GRBV. In this study, Tortistilus were collected on yellow panel cards across 102 vineyard sites and surrounding areas in Napa Valley, California, USA in 2021–2023. Specimens were morphotyped, sexed and tested for GRBV ingestion and acquisition by multiplex PCR or qPCR. Phylogenetic analysis of the partial sequence of mt-COI and ITS gene fragments of a subset of 40 Tortistilus specimens revealed clustering in a monophyletic clade with T. wickhami with the former barcode sequence. Only 6% (48/758) of the T. wickhami tested positive for GRBV, but none of the heads with salivary glands (0%, 0/50) of the dissected specimens tested positive for GRBV, indicating no virus acquisition. In contrast, half of the dissected heads with salivary glands of S. festinus (52%, 12/23), from the same collection vineyard sites, tested positive for GRBV. Together, our findings confirmed the presence of T. wickhami in northern California vineyards and suggested a dubious role of this treehopper as a vector of GRBV.

Changes in microbial communities during high-rate microbial selenate reduction in an up-flow anaerobic fluidized bed reactor

Biological fluidized bed reactor (FBR) is a promising treatment option for removing selenium oxyanions from wastewater by converting them into elemental selenium. The process can achieve high rates and be efficiently operated at low hydraulic retention times (HRT). However, the effects of HRT on the changes in microbial community in the FBR process have not been previously explored. In this study, dynamic changes of microbial communities both on biofilm carrier and in suspension of a selenate-reducing FBR were explored at various HRTs (0.3-120 h). Based on partial 16S rRNA gene sequencing of the microbial communities, alpha diversity of microbial communities in suspension rather than in the biofilm were impacted by low HRTs (0.3 h-3 h). Members from genera Geobacter, Geoalkalibacter, and Geovibrio were the main selenate-reducing bacteria on carrier throughout the FBR process. Genus Geobacter was dominant in FBR carrier at HRT of 24 h-120 h, whereas Geoalkalibacter and Geovibrio dominated at low HRT of 0.3 h-6 h. Suspended microbial communities detected in the FBR effluent were more sensitive to HRT changes than that in biofilm. “Shock loading” at HRT of 0.3 h had a great impact on microbial community compositions both in the biofilm and effluent. Reactor operation in batch mode and long HRT of 24 h helped to recover the community from “shock loading” and improved selenite reduction and ethanol oxidation. Redundancy analysis revealed that HRT, influent pH and selenate loading were key operational parameters that impacted both the FBR performance and the composition of microbial communities associated with both the FBR carrier and effluent. Overall, the microbial communities in FBR biofilm flexibly responded to the changes of HRT and showed resilience to the temporary shock loading, enabling efficient selenate removal.

Sustained Spread of HIV-1 CRF55_01B in Its Place of Initial Origin: Dynamics and Hotspots.

Shenzhen, a city with a substantial mobile population, was identified as the first discovered region of HIV-1 CRF55_01B and epicenter of its severe epidemic. During the implementation of venue-based behavioral interventions and the "treat-all" policy, discerning the spread patterns and transmission hotspots of CRF55_01B is imperative. In this study, 1,450 partial pol sequences, with demographic information, were collected from all newly diagnosed CRF55_01B infections in Shenzhen from 2008 to 2020. Molecular networks were constructed using the maximum likelihood and time-resolve phylogenies. Transmission rates, effective reproduction numbers (Re) of clusters and viral dispersal were evaluated using Bayesian inference. In total, 526 sequences formed 114 clusters, including seven large clusters. The status and size of clusters were strongly correlated with age, ethnicity, occupation and CD4+ T cell counts. The transmission rates of clusters were significantly higher than the national epidemic estimate. Four large clusters had Re exceeding 1 at the end of sampling period. Immigrants from Guangdong and Hunan, along with local residents, were identified as the transmission hubs, with heterosexual men being the main source and MSM being the main destination. The virus exhibited a high movement frequency from individuals aged 30-49 years toward diverse age groups. This study demonstrated the hidden CRF55_01B transmissions continued despite current combined interventions in Shenzhen, and special at-risk individuals susceptible to infection or transmission were identified, potentially serving as targets for more effective prevention and control of the local epidemic, thereby mitigating cross-regional spread nationwide due to population migration.

State-of-health estimation for lithium-ion batteries based on Kullback–Leibler divergence and a retentive network

Accurate state-of-health (SOH) estimation is crucial for the lithium-ion battery industry, as it underpins the safety, durability, and reliability of lithium-ion batteries. Currently, most researchers use various methods of health indicator (HI) extraction for the SOH estimation of batteries. However, these methods may require certain expertise and prior knowledge to achieve accurate modeling, being affected by measurement noise and other factors. To solve the abovementioned problems, three Kullback–Leibler (KL) divergence features based on partial voltage sequences are proposed as new HIs that are independent of prior knowledge and strongly correlated with SOH. Moreover, a modified retentive network is proposed to enhance SOH estimation accuracy and better utilize HIs than traditional deep learning methods, which have high training costs and insufficient accuracy. To ensure consistent extraction of KL divergence features across various experimental conditions and time intervals, a B-spline algorithm is utilized for interpolation. The effectiveness of the proposed method is validated through analysis of Pearson correlation coefficients and experiments conducted in four dimensions. Additionally, the potential of using the proposed method to compress data on the cloud-side is explored.

First Report of Klebsiella variicola Causing Tomato Pith Necrosis in Yunnan Province, China.

Tomato (Solanum lycopersicum L.) is a key vegetable crop in China. In August 2023, an outbreak of bacterial pith necrosis in tomato occurred in Lufeng County, Yunnan Province, China, affecting over 40% of the tomato plants in a greenhouse. The stems of infected plants developed a waterlogged soft rot and the disease progressed, the lower leaves and lateral branches of infected plants gradually wilted and died. A longitudinal cut of the stem revealed hollow pith with brown vascular tissue. To isolate the pathogen, the plant surface was disinfested with 75% ethanol. Then, a piece of infected tissue from the base of the stem was excised and immersed in sterile water for 2 min. A small amount of liquid was streaked onto TTC (2,3,5-triphenyltetrazolium chloride) agar medium using an inoculation loop, and plates were incubated at 28℃ for 24 h. Colonies on the TTC plate were white, indicating that the pathogen was not Ralstonia solanacearum. Colonies grown on LB (Luria-Bertani) agar medium were randomly selected and subjected to preliminary pathogenicity tests. Based on the results, a colony named Kv4 was selected and purified through six subcultures in LB agar medium. Biochemical tests showed the strain utilized D-sorbitol, raffinose and citrate but not adonitol, and was positive for methyl red, D-glucose (acid), urea hydrolysis, lysine decarboxylase, and motility, and negative for phenylalanine deaminase, H2S production, indole production, and ornithine decarboxylase. These characteristics align with Klebsiella species (Garrity et al. 2007). To determine the species of strain Kv4, partial sequences of the 16S rDNA, phoE, leuS, and rpoB genes were amplified (Barrios-Camacho et al. 2019) and sequenced. Through BLASTn analysis, strain Kv4 sequences of 16S rDNA (OR888750) had 99.47% identity (1488/1496 bp), phoE (OR899599) had 98.69% (605/613 bp) identity, leuS (OR899598) had 99.07% identity (959/968 bp), and rpoB (OR899597) had 97.69% (633/648 bp) identity with Klebsiella variicola strain FF0907. Using the ClustalW algorithm in MEGA11 for nucleotide sequence alignment, phylogenetic trees were constructed with 16S and phoE, leuS, and rpoB via the neighbor-joining method, confirming strain Kv4 as K. variicola. To test pathogenicity, the roots of 25 'Moneymaker' tomato plants with four to five true leaves were wounded, then each plant inoculated with a 15 mL bacterial suspension (OD600=0.6) of strain Kv4, while the control plants received sterile water. Plants were incubated at 28℃ with a 16 h photoperiod. Experiments were done twice. At 15 days after inoculation (DAI), all plants inoculated with Kv4 showed yellowing, unevenly distributed small black necrotic spots on the leaf surface, and purple-brown soft rot at the stem base. By 18 DAI, there was a gradual transformation of the stem bases from green to purplish brown. At 21 DAI, 60% of the inoculated plants displayed brownish soft rot at the stem base. In contrast, the control plants remained symptom-free. The pathogen was re-isolated from the stem and identified as K. variicola via sequence analysis of 16S, phoE, leuS, and rpoB. In recent years, several new bacterial pith necrosis diseases were reported in tomato (Guo et al. 2023; Ivić et al. 2023). This is the first study documenting K. variicola causing bacterial pith necrosis in tomato. Once considered a benign plant endophyte, Sun et al. (2023) reported K. variicola causing banana sheath rot in Guangdong and Guangxi Provinces, China. Malik et al. (2023) reported that K. variicola caused leaf streak in sorghum in India. This report of bacterial pith necrosis in tomato caused by K. variicola strain Kv4 underscores the escalating threat posed by emerging pathogens to agricultural production. The emergence of K. variicola as a tomato pathogen complicates plant disease management strategies.

Bactofencin YH, a novel bacteriocin with high inhibitory activity against clinical Streptococcus species.

Strain Lactiplantibacillus plantarum D1 with bacteriocin producing ability was found in the intestine of Gambusia affinis. The bacteriocin was found to have high inhibitory activity against multiple Streptococcus species and several other Gram-positive and Gram-negative bacteria. Bacteriocin was purified from culture supernatant by ion-exchange chromatography, Sep-Pak C18 cartridge, and reverse-phase high-performance liquid chromatography (RP-HPLC). Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectral analysis determined that purified bacteriocin has a molecular mass of 2,731 Da. A partial N-terminal sequence KRKKHKXQIYNNGM was obtained from the Edman analysis. The N-terminal sequence was employed to search against a translation of the draft genome of strain D1. The translated full amino acid sequence of the mature peptide is as follows: NH2- KRKKHKCQIYNNGMPTGQYRWC, which has a molecular weight of 2738 Da. A BLAST search revealed that this bacteriocin was most similar to bactofencin A but differed from it with three amino acid residues. No identical peptide or protein has been previously reported, and this peptide, termed bactofencin YH, was therefore considered to be a new bacteriocin produced by Lactiplantibacillus plantarum D1.

Analysis of the partial sequencing of clbA, clbB and clbQ in Escherichia coli isolates that produce colibactin and multilocus sequence typing

Colibactin, is a cyclomodulin expressed from polyketide synthase (pk) genomic islands. These bacterial toxins interfere with the eukaryotic cell cycle and induce DNA damage. The aim of the present study was to investigate the prevalence of colibactin production among E. coli strains recovered from different infections, determine the similarity of clb nucleotide sequences, and identify genotype of isolates using multilocus sequence typing(MLST). This was a prospective, cross-sectional study conducted from January 2022 to February 2023. A total of 117 clinical isolates were obtained from various sample types collected from outpatients and inpatients recruited to the Department of Bacteriology Labs in different hospitals in Baghdad, Iraq. clbA/clbR, clbB and clbP/clbQ were detected via conventional PCR, and partial sequencing of amplicons was performed via Sanger sequencing. For select isolates, MLST genotyping was performed. The most common phylogenetic group was B2 (61/106; 57.54%). Among the E. coli strains, 27/106 (25.47%) were clb + ve, and the most common type was clbB (13/27; 48.14%). Analysis of the partial sequencing of clb among the strains revealed high molecular similarity. Genotyping of 37 selected E. coli strains via MLST revealed 28 different genotypes. There was a high prevalence of colibactin production in phylogroup B2, and it seems that the clb + ve strains had conserved molecular structures. There was high genetic diversity among the strains tested.

Molecular Characterization and Gene Expression of Pattern Recognition Receptors in Brown-Marbled Grouper (Epinephelus fuscoguttatus) Fingerlings Responding to Vibriosis Infection

The pathogen recognition system involves receptors and genes that play a crucial role in activating innate immune response in brown-marbled grouper (Epinephelus fuscoguttatus) as a control agent against various infections including vibriosis. Here, we report the molecular cloning of partial open reading frames, sequences characterization, and expression profiles of Pattern Recognition Receptors (PRRs) in brown-marbled grouper. The PRRs, namely pglyrp5, tlr5, ctlD, and ctlE in brown-marbled grouper, possess conserved domains and showed shared evolutionary relationships with other fishes, humans, mammals, birds, reptilians, amphibians, and insects. In infection experiments, up to 50% mortality was found in brown-marbled grouper fingerlings infected with Vibrio alginolyticus compared to 27% mortality infected Vibrio parahaemolyticus and 100% survival of control groups. It is also demonstrated that all four PRRs had higher expression in samples infected with V. alginolyticus compared to V. parahaemolyticus. This PRRs gene expression analysis revealed that all four PRRs expressed rapidly at 4-h post-inoculation even though the Vibrio count was only detected earliest at 12-h post-inoculation in samples. The highest expression recorded was from V. alginolyticus inoculated fish spleen with up to 73-fold change for pglyrp5 gene, followed by 14 to 38-fold expression for the same treatment in spleen, head kidney, and blood samples for other PRRs, namely tlr5, ctlD, and ctlE genes. Meanwhile less than a 10% increase in expression of all four genes was detected in spleen, head kidney, and blood samples inoculated with V. parahaemolyticus. These findings indicated that pglyrp5, tlr5, ctlD, and ctlE play important roles in the early immune response to vibriosis infected, brown-marbled grouper fingerlings.

Reemergence of a Big Brown Bat Lyssavirus rabies Variant in Striped Skunks in Flagstaff, Arizona, USA, 2021-2023.

Background: Throughout the Americas, Lyssavirus rabies (RV) perpetuates as multiple variants among bat and mesocarnivore species. Interspecific RV spillover occurs on occasion, but clusters and viral host shifts are rare. The spillover and host shift of a big brown bat (Eptesicus fuscus) RV variant Ef-W1 into mesocarnivores was reported previously on several occasions during 2001-2009 in Flagstaff, Arizona, USA, and controlled through rabies vaccination of target wildlife. During autumn 2021, a new cluster of Ef-W1 RV cases infecting striped skunks (Mephitis mephitis) was detected from United States Department of Agriculture enhanced rabies surveillance in Flagstaff. The number of Ef-W1 RV spillover cases within a short timeframe suggested the potential for transmission between skunks and an emerging host shift. Materials and Methods: Whole and partial RV genomic sequencing was performed to evaluate the phylogenetic relationships of the 2021-2023 Ef-W1 cases infecting striped skunks with earlier outbreaks. Additionally, real-time reverse-transcriptase PCR (rtRT-PCR) was used to opportunistically compare viral RNA loads in brain and salivary gland tissues of naturally infected skunks. Results: Genomic RV sequencing revealed that the origin of the 2021-2023 epizootic of Ef-W1 RV was distinct from the multiple outbreaks detected from 2001-2009. Naturally infected skunks with the Ef-W1 RV showed greater viral RNA loads in the brain, but equivalent viral RNA loads in the mandibular salivary glands, compared to an opportunistic sample of skunks naturally infected with a South-Central skunk RV from northern Colorado, USA. Conclusion: Considering a high risk for onward transmission and spread of the Ef-W1 RV in Flagstaff, public outreach, enhanced rabies surveillance, and control efforts, focused on education, sample characterization, and vaccination, have been ongoing since 2021 to mitigate and prevent the spread and establishment of Ef-W1 RV in mesocarnivores.

Morphological and Phylogenetic Characterization of Trichomerium koreanum sp. nov. and Exophiala radicis Belonging to Chaetothyriales Fungi in Korea

The order Chaetothyriales encompasses black yeasts and related fungi known for their diverse morphologies, asexual reproduction, and habitat preferences. In this study, we isolated two fungal strains belonging to Chaetothyriales, designated KNUF-23-13A and KNUF-22-020, from a longhorn beetle (Cyrtoclytus capra) and soil sample, respectively, in Korea. Phylogenetic analyses based on the concatenated nucleotide sequences of the internal transcribed spacer regions and partial sequences of the large subunit (LSU) rRNA, β-tubulin (TUB2), and actin (ACT) genes revealed that the strains were grouped with strains of Trichomerium and Exophiala, respectively. For strain KNUF-23-13A, cultural and morphological differences were observed in the shape and size of mycelium between it and its phylogenetically closest species, Trichomerium lapideum CGMCC 3.17311T, supporting the novelty of this isolated species. In the case of strain KNUF-22-020, the cultural and morphological characteristics were similar to those previously reported for E. radicis P2854T. Based on morphological and phylogenetic analyses, we revealed KNUF-23-13A as a novel species, which we describe and name T. koreanum sp. nov., and KNUF-22-020 as Exophiala radicis, a previously unreported species in Korea.

First report of Tomato interveinal chlorosis virus infecting Rhynchosia minima plants in Brazil.

Tomato interveinal chlorosis virus (ToICV; Begomovirus solanumintervenae, genus Begomovirus, family Geminiviridae) has been described infecting tomato (Solanum lycopersicum) and Macroptilium lathyroides in Northeastern (NE) Brazil for more than a decade (Albuquerque et al., 2012; Silva et al., 2012). During a survey in 2020, plants of the leguminous weed Rhynchosia minima exhibiting virus-like symptoms such as mosaic and interveinal chlorosis were observed in the state of Alagoas, NE Brazil. Symptomatic leaf samples of R. minima were randomly collected (n=15; supplementary figure 1). Total DNA from each sample was used as a template for PCR amplification of partial begomoviral DNA-A sequences using the degenerate primer pair PAL1v1978 and PAR1c496, universal for geminiviruses (Rojas et al., 1993). Amplicons of ~1.2 kbp were observed from 12 samples, although this should not be considered as incidence since only symptomatic plants were collected. To identify the begomovirus associated with R. minima, viral genomes were amplified from PCR-positive samples using rolling circle amplification (RCA) (Inoue-Nagata et al., 2004). The RCA products were digested with HindIII, cloned into the pBluescript II KS+ plasmid vector and bidirectionally Sanger-sequenced (Macrogen Inc., Seoul). BLASTn searches indicated that the clones (n=4) reported here corresponded to a begomovirus DNA-A component, and pairwise comparisons showed that they shared the highest identity with ToICV, at 92.4-94.7% nucleotide sequence identity. Based on the species demarcation criteria of ≥91% nucleotide identity for the genus Begomovirus (Brown et al., 2015), the begomoviruses obtained from R. minima are new isolates of ToICV. The new DNA-A sequences of 2,619-2,623 nt in length were deposited in GenBank under accession numbers PP639092 to PP639095. Multiple nucleotide sequence alignments were prepared using the MUSCLE algorithm implemented in MEGA v.11 (Kumar et al., 2018), and a maximum likelihood (ML) tree was reconstructed in RaxML-NG (Kozlov et al., 2019), assuming a general time reversible (GTR) nucleotide substitution model with a gamma (G) model of rate heterogeneity and 1,000 bootstrap replicates. The DNA-A-based tree showed that the ToICV sequences clustered into a monophyletic group, additionally supporting these isolates as members of the species Begomovirus solanumintervenae. At least two independent interspecies recombination events were predicted among the ToICV isolates, with breakpoints located in the Rep-encoding region and ToICV (GenBank Accession JF803253), tomato mottle leaf curl virus (JF803248) and soybean blistering mosaic virus (MN486865) detected as putative parents. To the best of our knowledge, this is the first report of ToICV infecting R. minima worldwide, expanding the host range of this begomovirus. Non-cultivated plants such as R. minima play a crucial role as reservoirs and sources of inoculum for begomoviruses (Paz-Carrasco et al., 2014), reinforcing their relevance to socioeconomically important crops.

Identification and Full-Length Sequence Analysis of a Novel Recombinant Goat Astrovirus Genotype in Guangxi, China.

Astroviruses are single-stranded, positive-sense RNA viruses capable of infecting humans as well as a wide range of mammalian and avian species, with a length of approximately 6.6-7.7 kb. In this study, 139 goat fecal samples collected from the Guangxi province were used for the RT-PCR detection, and two of these were positive for goat astrovirus, with a positivity rate of 1.44% (2/139). The complete genome sequence of an astrovirus strain and the partial genome sequence of a strain astrovirus, named GX WZ 2023 and GX HC 2023, were amplified and sequenced, and their sequence lengths were 6284 nt and 6213 nt, respectively. Among them, the capsid protein of goat astrovirus GX HC 2023 showed the highest amino acid identity of 95.9% with ovine astrovirus GX, which belonged to the MAstV-2 genotype. However, the closest relative of the GX WZ 2023 strain was found to be the caprine astrovirus Sichuan, with a nucleotide sequence identity of 76.8%. The ORF1ab nonstructural protein of this strain showed the highest amino acid identities of 89.2 and 95.8% with the ovine astrovirus S5.1 and caprine astrovirus G5.1 strains, respectively. However, its ORF2 capsid protein has 68.4% amino acid identity with the bovine astrovirus (BAstV) 16 2021 CHN strain and only 21.9-64% amino acid identity with all available strains of goat astrovirus. The GX WZ 2023 strain was recombined with the Chinese (BAstV 16 2021 CHN) and Japanese bovine strains (BAstV JPN 2015) in the ORF2 region. Therefore, the goat astrovirus GX WZ 2023 is proposed as a new member of the family goat astroviridae based on the species classification criteria of the International Committee on Taxonomy of Viruses. These findings enhance our understanding of the prevalence and genetic evolution of goat astrovirus and provide a scientific basis for future studies of these viruses in other animals.

Prevalence and Sequence Analysis of Equine Rhinitis Viruses among Horses in Poland.

Equine rhinitis A (ERAV) and B (ERBV) viruses are respiratory pathogens with worldwide distribution. The current study aimed to determine the frequency of infection of ERAV and ERBV among horses and foals at Polish national studs, and to determine genetic variability within the viruses obtained. Virus-specific quantitative RT-PCR assays targeting a 5' untranslated region were used to screen nasal swabs collected from 621 horses at 16 national horse studs from throughout Poland, including 553 healthy horses and 68 horses with respiratory disease. A partial DNA polymerase gene was amplified and sequenced from the qRT-PCR-positive samples. The obtained sequences were analysed using phylogeny and genetic network analysis. None of the nasal swabs were positive for ERAV, whereas ERBV was found in 11/621 (1.78%) samples collected from 10 healthy horses and one foal affected by respiratory disease. Partial DNA polymerase gene sequence variability was correlated with individual horses and studs from which samples were collected when only Polish sequences were analysed, but there was no correlation between country of origin and ERBV sequence when Polish and international sequences were included in the network. The report presents the first detection of ERBV in Poland.

Horse Meat Microbiota: Determination of Biofilm Formation and Antibiotic Resistance of Isolated Staphylococcus Spp.

Domestic horses could be bred for leisure activities and meat production, as is already the case in many countries. Horse meat is consumed in various countries, including Kazakhstan and Kyrgyzstan, and with the increase in this consumption, horses are registered as livestock by the Food and Agricultural Organization. In this study, horse meat microbiota of horse samples (n = 56; 32 samples from Kazakhstan and 24 samples from Kyrgyzstan) from two countries, Kazakhstan (n = 3) and Kyrgyzstan (n = 1), were investigated for the first time by next-generation sequencing and metabarcoding analysis. The results demonstrated that Firmicutes, Proteobacteria, and Actinobacteria were the dominant bacterial phyla in all samples. In addition, three (5.4%) Staphylococcus strains were isolated from the Uzynagash region, Kazakhstan. Staphylococcus strains were identified as Staphylococcus warneri, S. epidermidis, and S. pasteuri by partial 16S rRNA DNA gene Sanger sequencing. All three Staphylococcus isolates were nonbiofilm formers; only the S. pasteuri was detected as multidrug-resistant (resistant to penicillin, cefoxitin, and oxacillin). In addition, S. pasteuri was found to carry mecA, mecC, and tetK genes. This is the first study to detect potentially pathogenic Staphylococcus spp. in horse meat samples originating from Kazakhstan. In conclusion, it should be carefully considered that undercooked horse meat may pose a risk to consumers in terms of pathogens such as antibiotic-resistant Staphylococcus isolates.

Exploring Listeria monocytogenes in Ewe Milk: ssrA Gene-based Real-time PCR Identification, Phylogenetic Analysis, and Antibacterial Assessment of Magnesium Oxide Nanoparticles Synthesized with Myrtus communis Leaf Extract

Background: Listeria monocytogenes, a zoonotic pathogen affecting humans and animals, exhibits a global distribution, including Iraq. This study focused on the rapid identification and phylogenetic analysis of L. monocytogenes in freshly collected ewe milk samples from various farms in Al-Qadisiyah province, Iraq.Methods: The study was conducted with care and precision, involving 150 milk samples. These samples were subjected to traditional bacterial isolation and identification using the enrichment culture method and biochemical tests, with the PCR technique confirming the results. The antibacterial activity of MgONPs was then assessed using the disc diffusion method, ensuring a comprehensive and reliable approach to the study.Result: The results show that 150 ewe milk samples underwent real-time PCR (RT-PCR) targeting the ssrA gene, followed by partial 16S rRNA gene sequencing (PSGS) of purified conventional PCR products. Furthermore, the study entails the biosynthesis of magnesium oxide nanoparticles using Myrtus communis leaf extract, followed by a comprehensive characterization utilizing UV-spectra, FTIR, SEM, and TEM techniques. The Agar well diffusion method assessed the antibacterial efficacy of these Biosynthesized nanoparticles against L. monocytogenes. The RT-PCR results revealed the presence of L. monocytogenes in 36 out of 150 samples (24%). Subsequent PCR analysis confirmed the presence of the pathogen in 30 out of these 36 positive samples (83.33%). Sequencing of two purified PCR products demonstrated 100% nucleotide identity with global isolates from Iraq and Turkey. Furthermore, the study demonstrated that L. monocytogenes exhibited substantial sensitivity (24.66 ± 0.3) to the biosynthesized magnesium oxide nanoparticles. These findings underscore the speed and precision of the RT-PCR method for detecting L. monocytogenes in fresh ewe milk samples.Conclusion: This comprehensive investigation enhances our understanding of L. monocytogenes prevalence in ewe milk and highlights the potential of Myrtus communis -derived nanoparticles for combating this pathogen.Keywords: Antibacterial nanoparticles; Listeria monocytogenes; Magnesium oxide; Myrtus communis sheep milk; ssrA gene

Lure specificity, phenology, and damage caused by Epiphyas moths (Lepidoptera: Tortricidae) in Western Australian apple orchards.

Multiple Epiphyas species inhabit southwestern Western Australia, including Light Brown Apple Moth (LBAM) Epiphyas postvittana (Walker) (Lepidoptera: Tortricidae), a globally significant, polyphagous pest. This study evaluated the efficacy and specificity of lures designed for 3 Epiphyas species: E. postvittana, Epiphyas pulla (Turner), and the undescribed Epiphyas sp. (1) (Common). Additionally, the study sought to determine the presence and distribution of Epiphyas species in 3 significant apple-growing localities. Trapping, together with partial sequencing of the mitochondrial COI gene, found LBAM to be restricted to the Perth Hills and E. pulla, to apple orchards near Manjimup and Pemberton. This geographic disjunction remains unexplained. Epiphyas sp. (1) was not recorded despite using a specifically designed lure. The E. pulla and LBAM traps demonstrated superior efficacy in capturing their target species, while the catch in Epiphyas sp. (1) traps did not significantly differ between the 2. Both E. pulla and LBAM exhibited peak abundance from late spring to the end of summer (October-February), with variations in timing and peak catch of male moths across species, locations, and years. Surveys conducted in April during the harvest period (February-May), when moth traps caught an average of 1-1.8 moths/trap/week, found no Epiphyas larvae or damage on 140,400 mature apples or on 26,000 leaves. While E. pulla and LBAM traps effectively monitor their target moths, genetic identification of trap catch would be necessary if they co-occurred. Encouragingly, the results indicate that both species become relatively rare as harvest season approaches, and neither inflicts significant damage to mature apples under existing management.

Molecular detection and identification of Enteromonas species in human and animal hosts using polymerase chain reaction and DNA sequencing

Enteromonas hominis, a human intestinal protozoan parasite of the diplomonad group, has been overlooked because of its commensal features; therefore, molecular studies on this parasite are limited. To address this gap, we designed a molecular screening protocol using polymerase chain reaction (PCR) and DNA sequencing targeting the 18S small subunit ribosomal RNA gene and applied this screening method to the molecular epidemiological analysis of Enteromonas spp. in humans and various livestock. We validated our methodology using stool samples collected from 215 humans and 270 animal hosts (buffaloes, pigs, dogs, goats, horses, rodents, chickens, and ducks) during an annual epidemiological investigation conducted from 2013 to 2016 on Sumba Island, Indonesia. The overall prevalences of Enteromonas spp. were 33.9 % (n = 73/215) in humans and 25.2 % (n = 68/270) in mammals and avians. The positive predictive value of this PCR method for Enteromonas spp., as evaluated through sequencing, was 90.1 % in human samples and 58.1 % in non-human samples (particularly low, 11.4 % in rodents). Although the specificity of the PCR approach may not be perfect, in combination with DNA sequencing, it was effective in detecting and identifying a partial sequence (1458 bp) of the target gene region in Enteromonas species.

Morphological and molecular data on the bat flies, Basilia silvae (Brèthes, 1913) and Trichobius parasiticus Gervais, 1844 parasitizing Chilean bats.

Among mammals, bats harbour the greatest taxonomic diversity of ectoparasitic arthropods. This is mainly due to their high mobility, wide distribution range and gregarious social behaviour. In Chile, 17 species of bats have been reported; however, their ectoparasitic arthropofauna has been little studied. There are currently 12 taxa recorded, belonging to two classes and associated with only four species of bats. This study aimed to investigate the diversity of parasitic flies associated with bats in three ecoregions: Atacama Desert, Chilean Matorral and Valdivian temperate forest. During 2021, using mist nets, bats were captured in Anzota caves (Atacama Desert ecoregion), Huelquén and Alto Jahuel (Chilean Matorral ecoregion), Cherquenco and San Patricio (Valdivian temperate forest). Each bat was carefully checked during a 15-min interval for the collection of ectoparasites. The arthropods were deposited in vials with 96% ethanol. A total of 26 bats corresponding to three species (Vespertilionidae: Histiotus magellanicus Philippi, 1866, Myotis arescens (Osgood, 1943); Phyllostomidae: Desmodus rotundus (Geoffroy, 1810)) were captured from which a total of 142 ectoparasitic arthropods were collected. Bat flies were separated/identified under a stereomicroscope. Additionally, from the fieldwork, we report the presence of other ectoparasites associated with Chilean bats. In our study, we report new host-parasite associations between Trichobius parasiticus Gervais, 1844 (Diptera: Streblidae) on D. rotundus, and Basilia silvae (Brèthes, 1913) (Diptera: Nycteribiidae) in M. arescens in Chile. Our study extended the latitudinal range of distribution for B. silvae to Araucanía region, and we report for first time T. parasiticus in the country. Additionally, partial sequences of the cytochrome c oxidase I gene were obtained from these specimens. Although there is slight morphological variation in the specimens of T. parasiticus, phylogenetic analyses suggest that they correspond to the same species. The sequences generated for B. silvae represent the first for the species. Authors recommend the use of an integrative approach in the identification of ectoparasites in poorly studied ecoregions and hosts. The integration of different markers is necessary to determine more precisely the phylogenetic relationships between South American populations and species of the genera Basilia and Trichobius.