Complete Genome Sequence Analysis Research Articles

Novel Intertypic Recombinant Coxsackievirus A2 Containing Specific Amino Acid Mutations in the RNA-Dependent RNA Polymerase Potentially Associated With Its Emergence.

Coxsackievirus A2 (CVA2), a member of enterovirus A species (EV-A), is associated with diverse human diseases and occasionally causes acute gastroenteritis (AGE). In Thailand, CVA2 emerged as the predominant genotype in 2019. The increasing incidence of CVA2, coupled with the limited availability of full-length genomes, highlights the need for more complete genome sequence analysis to facilitate molecular epidemiology study. This study aimed to investigate the molecular epidemiology, evolutionary dynamics, and recombination characteristics of CVA2 associated with AGE in Thailand from 2013 to 2022. A total of 19 full-genome sequences of CVA2 isolated from stool samples of AGE patients in Thailand were characterized and analyzed together with the reference sequences available in the GenBank database. A novel lineage of CVA2 (subgenotype C5) was detected with the potential recombination with CVA10 within the P2 and P3 regions. Specific consensus amino acid mutations, A61S in the VP3 gene and R136K in the 3D (RdRp) gene, were identified in all CVA2 recombinant strains. Additionally, the S45G mutation in the RdRp gene was found to be potentially associated with the emergence of CVA2 infection in 2019. In conclusion, this study reveals potential intertypic recombinant events and specific mutations in CVA2 strains isolated from AGE patients and provides a broader understanding of its evolutionary epidemiology.

Complete genome sequence analysis and Pks genes identification of Brevibacillus brevis FJAT-0809-GLX with a broad inhibitory spectrum against phytopathogens.

Brevibacillus brevis FJAT-0809-GLX has a broad spectrum of antimicrobial activity. Understanding the molecular basis of biocontrol ability of B. brevis will allow us to develop effective microbial agents for sustainable agriculture. In this study, we present the complete and annotated genome sequence of FJAT-0809-GLX. The complete genome size of B. brevis FJAT-0809-GLX was 6,137,019bp, with 5688 predicted coding sequences (CDS). The average GC content of 47.38%, and there were 44 copies of the rRNAs operon (16S, 23S and 5S RNA), and 127 tRNA genes. A total of 11,162 genes were functionally annotated with the COG, GO, and KEGG databases, and 123 genes belonged to CAZymes. Genomic secondary metabolite analysis indicated 13 clusters encoding potential new antimicrobials. FJAT-0809-GLX was designated as B. brevis according to average nucleotide polymorphism (ANI) and phylogenetic analysis. The pangenome consisted of 7141 homologous genes, and 4469 homologous genes shared by B. brevis FJAT-0809-GLX, B. brevis NBRC100599, B. brevis DSM30, and B. brevis NCTC2611. The number of unique homologous genes of B. brevis FJAT-0809-GLX (419 genes) and B. brevis NBRC100599 (480 genes) were much more than those in B. brevis DSM30 (13 genes), and B. brevis NCTC2611 (6 genes). Nine gene clusters encoding for secondary metabolite biosynthesis were compared in the genome of B. brevis FJAT-0809-GLX with those of B. brevis NBRC100599, B. brevis DSM30 and B. brevis NCTC2611, and the gene clusters encoding for lantipeptide and transatpks-otherks only existed in genome of B. brevis FJAT-0809-GLX. The 11 BbPks genes were included in the B. brevis FJAT-0809-GLX genome, which contained the conserved PS-DH domain. The relative expression of BbPksL, BbPksM2, BbPksM3, BbPksN3, BbPksN4 and BbPksN5 reached a maximum at 120h and then decreased at 144h. Our results provided detailed genomic and Pks genes information for the FJAT-0809-GLX strain, and lid a foundation for studying its biocontrol mechanisms.

Whole-Genome Sequencing of Newly Emerged Fungal Pathogen Aspergillus Lentulus and Its Azole Resistance Gene Prediction.

Aims: Aspergillus lentulus is an important newly recorded species in the A. fumigatus complex and its resistance to azole drugs and the high mortality rate of infected individuals have emerged as problems. Comprehensive understanding of the A. lentulus is limited due to lack of genome-wide fine mapping data. The aim of this study was to investigate the A. lentulus signature at the molecular level, analyze the genome-wide profile of this strain, and predict its possible genes that execute azole resistance. Methods: In this study, a whole-genome sequencing of a clinically isolated A. lentulus strain (named A. lentulus PWCAL1) was studied by PacBio Sequel sequencing platform. Azole resistance genes were predicted based on whole-genome sequencing data analysis, gene function annotation, comparative genomic analysis, and BLASTP alignment using the Mycology Antifungal Resistance Database to comprehensively understanding the genome-wide features, pathogenicity, and resistance mechanisms of A. lentulus. Results: The results of whole-genome sequencing demonstrated that the total length of A. lentulus PWCAL1 genome was 31255105 bp, the GC content was 49.24%, and 6883 coding genes were predicted. A total of 4565, 1824, and 6405 genes were annotated in the Gene Ontology, Clusters of Orthologous Groups, and Kyoto Encyclopedia of Genes and Genomes databases, respectively. In the Pathogen Host Interactions Database and the Database of Fungal Virulence Factors, 949 and 259 interacting virulence factors were identified, respectively, with the main virulence factor-mutant virulence phenotype, being enriched in reduced virulence. Comparative genomic analysis showed that there were 5456 consensus core genes in this strain and four closely related strains of A. fumigatus complex, which were mainly involved in human diseases, metabolism, organismal systems, etc. Among the three aligned A. lentulus strains, the number of unique genes of this bacterium was the highest with a number of 171, and these genes were mainly associated with carbohydrate metabolism and cell growth and death. Resistance gene prediction demonstrated that the A5653 gene of this bacterium had F46Y/N248T double point mutations on the CYP51A gene, but no tandem repeat mutations in the promoter region were detected. Furthermore, 12 genes belonging to the fungal multidrug resistance ATP-binding cassette (ABC) transporters were identified based on the complete genome sequence and phylogenetic analysis of A. lentulus, which belonged to the ALDp subfamily, the PDR subfamily (AtrB, CDR1, and CDR2), and the MDR subfamily (MDR1), respectively, and there were four genes that are annotated to the major facilitator superfamily multidrug transporter. Further phylogenetic tree classification of the ABC transporter subfamilies predicted in the nine selected A. fumigatus complex strains showed that these putative ABC proteins were divided into two main clusters, which belonged to the PDR (CDR1, CDR2, AtrB, and AtrF) and MDR subfamilies (MDR1, MDR2, and MDR3). The distribution of these ABC proteins varies among different species of the A. fumigatus complex. Conclusions: The main result obtained from this study for the whole genome of A. lentulus provide new insights into better understanding the biological characteristics, pathogenicity, and resistance mechanisms of this bacterium. In this study, two resistance mechanisms, which include CYP51A gene mutation and multidrug-resistant ABC transporter, were predicted in a single isolate. Based on the predicted CYP51A-F46Y/N248T site mutation combination, we speculate that the CYP51A gene of A. lentulus may be partially responsible for azole resistance. Based on the predicted ABC transporter family genes, we hypothesize that resistance to multiple azoles in A. lentulus is mediated, at least in part, by these ABC transporters with resistance.

Findings of tobacco ringspot virus in ornamentals in the Netherlands from 1997 to 2020 indicate a need for evaluation of its European Union quarantine status

AbstractOccasional findings of tobacco ringspot virus (TRSV) in the Netherlands since 1997, were the reason for large-scale surveys for TRSV in vegetatively propagated ornamental plants and for its vector, Xiphinema americanum sensu lato in soil. TRSV was identified in thirteen ornamental species, comprising over 30 cultivars, while X. americanum s.l. was not found. This report also describes a newly designed and validated real-time RT-PCR and includes phylogenetic analyses of (near) complete genome sequences from available TRSV isolates and NCBI GenBank accessions in relation to their metadata of host and origin. Overall, the results suggest that TRSV entered the Netherlands on multiple occasions and was further spread by vegetative propagation. Furthermore, TRSV seems to be more widespread in the European Union (EU) than previously assumed, indicating that the current regulatory measures do not guarantee the absence of TRSV. Alternative strategies to protect the crops at risk are discussed, and reconsideration of the regulatory status of TRSV in the EU is recommended.

Complete genome sequence analysis of Pestalotiopsis microspora, a fungal pathogen causing kiwifruit postharvest rots

BackgroundThe postharvest rot of kiwifruit is one of the most devastating diseases affecting kiwifruit quality worldwide. However, the genomic basis and pathogenicity mechanisms of kiwifruit rot pathogens are lacking. Here we report the first whole genome sequence of Pestalotiopsis microspora, one of the main pathogens causing postharvest kiwifruit rot in China. The genome of strain KFRD-2 was sequenced, de novo assembled, and analyzed.ResultsThe genome of KFRD-2 was estimated to be approximately 50.31 Mb in size, with an overall GC content of 50.25%. Among 14,711 predicted genes, 14,423 (98.04%) exhibited significant matches to genes in the NCBI nr database. A phylogenetic analysis of 26 known pathogenic fungi, including P. microspora KFRD-2, based on conserved orthologous genes, revealed that KFRD-2’s closest evolutionary relationships were to Neopestalotiopsis spp. Among KFRD-2’s coding genes, 870 putative CAZy genes spanned six classes of CAZys, which play roles in degrading plant cell walls. Out of the 25 other plant pathogenic fungi, P. microspora possessed a greater number of CAZy genes than 22 and was especially enriched in GH and AA genes. A total of 845 transcription factors and 86 secondary metabolism gene clusters were predicted, representing various types. Furthermore, 28 effectors and 109 virulence-enhanced factors were identified using the PHI (pathogen host-interacting) database.ConclusionThis complete genome sequence analysis of the kiwifruit postharvest rot pathogen P. microspora enriches our understanding its disease pathogenesis and virulence. This study establishes a theoretical foundation for future investigations into the pathogenic mechanisms of P. microspora and the development of enhanced strategies for the efficient management of kiwifruit postharvest rots.

Complete genome sequence of a novel mitovirus isolated from the phytopathogenic fungus Alternaria alternata causing apple leaf blotch.

In this study, a novel mitovirus, tentatively designated as "Alternaria alternata mitovirus 2" (AaMV2), was isolated from the fungus Alternaria alternata f. sp. mali causing apple leaf blotch disease. The complete genome of AaMV2 is 3,157 nucleotides in length, with an A+U content of 68.10%. The genome has a single large open reading frame (ORF) encoding an RNA-dependent RNA polymerase (RdRp) protein with a molecular mass of 98.10 kDa. BLAST analysis revealed that AaMV2 has the highest sequence identity to Leptosphaeria biglobosa mitovirus 6, with 79.76% and 82.86% identity at the amino acid and nucleotide level, respectively. Phylogenetic analysis suggested that AaMV2 is a new member of the genus Duamitovirus within the family Mitoviridae. This is the first report of the complete genome sequence analysis of a mitovirus in A. alternata.

Complete genome sequence analysis of Reticuloendotheliosis virus integrated in nonhomologous Avipoxvirus

Integration of nucleic acid sequences of Reticuloendotheliosis virus (REV) in Avipoxvirus（APV） has become commonplace. In this study, 4 strains of suspected Fowlpox virus (FPV) and 1 strain of suspected Pigeonpox virus (PPV) collected in Taiyuan, Shanxi Province were cultured in chicken embryos, and the 4b core protein gene was amplified by PCR, and the identity and genome similarity were determined by sequence analysis. The sequences between the end of ORF201 and the beginning of ORF203 of FPV and PPV were then amplified, sequenced, and subjected to sequence comparison to determine genome similarity. The results showed that the isolates were 4 strains of FPV and 1 strain of PPV. The 4 isolated strains of FPV belong to type A1 virus, with 100 % identity to each other and to the FWPV-09-Jilin strain isolated in Jilin, China, and the lowest identity to the type B2 virus TNPV5/NZL/2009, which is only 74 %. PPV belongs to type A2 virus, and its identity with local strain of fowlpox virus was 90.1 %, with the highest identity of 100 % with PPLH and ROPI/W370/ON/2012 and ow_2017_3 strains, which also belong to type A2 pigeonpox virus, and the lowest identity of 73.7 % with TNPV5/NZL/2009, a type B2 virus. The complete genome of REV sequences integrated into FPV and PPV were amplified, and 5 REV nucleic acid sequences were obtained after sequencing and concatenation, with lengths ranging from 7942 to 8005 bp. The identity analysis results indicate that it has high identity with isolates from Northeast China, Guangdong, and Guangxi regions in China. Based on its gp90 protein gene, the REV integrated into the poxvirus belong to type III, with the highest identity of 99.9% with strains such as APC-566 and CY1111, and the lowest identity with REV-Anhui1, at 95.4 %. The length of the pol gene varies among different strains of REV, and its encoded amino acid changes significantly after position 675, with deletions and alterations. This study indicates that all fowlpox viruses isolated in Taiyuan, Shanxi Province have integrated the entire REV gene sequence, with high identity between them. At the same time, it indicates that the pigeonpox virus isolate has also integrated the entire REV gene sequence, and has the highest identity with the integrated REV gene sequence in fowlpox virus.

Complete genome sequence, phenotypic correlation and pangenome analysis of uropathogenic Klebsiella spp

Urinary tract infections (UTI) by antibiotic resistant and virulent K. pneumoniae are a growing concern. Understanding the genome and validating the genomic profile along with pangenome analysis will facilitate surveillance of high-risk clones of K. pneumoniae to underpin management strategies toward early detection. The present study aims to correlate resistome with phenotypic antimicrobial resistance and virulome with pathogenicity in Klebsiella spp. The present study aimed to perform complete genome sequences of Klebsiella spp. and to analyse the correlation of resistome with phenotypic antimicrobial resistance and virulome with pathogenicity. To understand the resistome, pangenome and virulome in the Klebsiella spp, the ResFinder, CARD, IS Finder, PlasmidFinder, PHASTER, Roary, VFDB were used. The phenotypic susceptibility profiling identified the uropathogenic kp3 to exhibit multi drug resistance. The resistome and in vitro antimicrobial profiling showed concordance with all the tested antibiotics against the study strains. Hypermucoviscosity was not observed for any of the test isolates; this phenotypic character matches perfectly with the absence of rmpA and magA genes. To the best of our knowledge, this is the first report on the presence of ste, stf, stc and sti major fimbrial operons of Salmonella enterica serotype Typhimurium in K. pneumoniae genome. The study identifies the discordance of virulome and virulence in Klebsiella spp. The complete genome analysis and phenotypic correlation identify uropathogenic K. pneumoniae kp3 as a carbapenem-resistant and virulent pathogen. The Pangenome of K. pneumoniae was open suggesting high genetic diversity. Diverse K serotypes were observed. Sequence typing reveals the prevalence of K. pneumoniae high-risk clones in UTI catheterised patients. The study also highlights the concordance of resistome and in vitro susceptibility tests. Importantly, the study identifies the necessity of virulome and phenotypic virulence markers for timely diagnosis and immediate treatment for the management of high-risk K. pneumoniae clones.

Isolation and complete genome sequence analysis of Vibrio rotiferianus strain TO-01 from Trachinotus ovatus to reveal its pathogenicity and drug resistance

Vibrio rotiferianus is a pathogenic bacterium that is widely distributed in the ocean and can cause diseases in a variety of marine animals, including fish, shrimp and shellfish. In this study, we isolated and identified V. rotiferianus strain TO-01 from diseased Trachinotus ovatus. Infected fish exhibited a range of pathological symptoms, including lethargy in swimming, blackened body coloration, skin ulceration and erratic side swimming. Tissue section examination revealed pronounced damage to the liver, spleen and gills of the infected fish. An analysis of immune factor expression revealed a significant increase in the expression of inflammatory factors, including interleukin (IL)-1, IL-2, IL-6 receptor and IL-21, in the spleen and kidney of fish infected with TO-01 at 6 h post-infection. To gain a more in-depth comprehension of the pathogenicity and drug resistance of V. rotiferianus, we conducted whole genome sequencing. The total genome length of V. rotiferianus was 4734,075 bp, and 74 virulence factors were predicted within the genome. Most of these virulence genes were associated with bacterial adhesion and flagellar motility. A gene associated with group sensing, cqsA, and a gene associated with hemolysin, tlh, were also identified. The prediction of drug resistance genes was in agreement with the results of drug sensitivity assays, both of which indicated that TO-01 has significant resistance to β-lactams. The findings of this study significantly enhance our understanding of the pathogenic mechanism of V. rotiferianus and give crucial insights into disease prevention and control in the management of T. ovatus.

Transfection of a Molecular Clone of Naegleria gruberi rDNA into N. gruberi Trophozoites.

All ribosomal genes of Naegleria trophozoites are maintained in a closed circular extrachromosomal ribosomal DNA (rDNA) containing element (CERE). While little is known about the CERE, a complete genome sequence analysis of three Naegleria species clearly demonstrates that there are no rDNA cistrons in the nuclear genome. Furthermore, a single DNA origin of replication has been mapped in the N. gruberi CERE, supporting the hypothesis that CERE replicates independently of the nuclear genome. This CERE characteristic suggests that it may be possible to use engineered CERE to introduce foreign proteins into Naegleria trophozoites. As the first step in exploring the use of a CERE as a vector in Naegleria, we developed a protocol to transfect N. gruberi with a molecular clone of the N. gruberi CERE cloned into pGEM7zf+ (pGRUB). Following transfection, pGRUB was readily detected in N. gruberi trophozoites for at least seven passages, as well as through encystment and excystment. As a control, trophozoites were transfected with the backbone vector, pGEM7zf+, without the N. gruberi sequences (pGEM). pGEM was not detected after the first passage following transfection into N. gruberi, indicating its inability to replicate in a eukaryotic organism. These studies describe a transfection protocol for Naegleria trophozoites and demonstrate that the bacterial plasmid sequence in pGRUB does not inhibit successful transfection and replication of the transfected CERE clone. Furthermore, this transfection protocol will be critical in understanding the minimal sequence of the CERE that drives its replication in trophozoites, as well as identifying regulatory regions in the non-ribosomal sequence (NRS).

Complete genome sequence and phylogenetic analysis of a newly discovered fusagra-like virus infecting Carica papaya in Ecuador.

A new fusagra-like virus infecting papaya (Carica papaya L.) was genetically characterized. The genome of the virus, provisionally named "papaya sticky fruit-associated virus" (PSFaV), is a single molecule of double-stranded RNA, 9,199 nucleotides (nt) in length, containing two discontinuous open reading frames. Pairwise sequence comparisons based on complete RNA-dependent-RNA-polymerase (RdRp) sequences revealed identity of 79.4% and 83.3% at the nt and amino acid (aa) level, respectively, to babaco meleira-like virus (BabMelV), an uncharacterized virus sequence discovered in babaco (Vasconcellea x heilbornii) in Ecuador. Additional plant-associated viruses with sequence identity in the 50% range included papaya meleira virus (PMeV) isolates from Brazil. Phylogenetic analysis based on the amino acid sequences of the capsid protein (CP), RdRp, and CP-RdRp fusion protein genes placed PSFaV in a group within a well-supported clade that shares a recent ancestor with Sclerotium rolfsii RNA virus 2 and Phlebiopsis gigantea mycovirus dsRNA 2, two fungus-associated fusagraviruses. Genomic features and phylogenetic relatedness suggest that PSFaV, along with its closest relative BabMelV, represent a species of novel plant-associated virus classified within the recently established family Fusagraviridae.

Integrative genomic and transcriptomic analysis of Xanthomonas oryzae pv. oryzae pathotype IV, V, and IX in China reveals rice defense-responsive genes

Bacterial blight of rice is a devastating disease caused by the gram-negative bacteria Xanthomonas oryzae pv. oryzae (Xoo). Chinese Xoo strain pathotypes IV, V, and IX are the major virulent Xoo strain types in South China sequentially from the 1990s to the present. Here, we report the isolation of GD0201 and GD0202, which belong to pathotypes IV and IX, respectively, and the complete genome sequence and transcriptomic analysis of GD0201 (IV), GD1358 (V), and GD0202 (IX). We found that resistance genes xa5, Xa23, and Xa27 confer strong resistance to all three Xoo strains, indicating that they are currently good choices for resistance rice breeding. The genome analysis reveals fewer TAL and non-TAL effector coding genes in GD0202 than in the other two strains, potentially contributing to its strong virulence. Transcriptomic analysis of ZH11 inoculated with the three Xoo strains strongly suggests that three Xoo strains for better infection repress the ethylene response factor (ERF) gene family members. Furthermore, weighted gene co-expression network analysis (WGCNA) and protein-protein interaction (PPI) analysis revealed 14 hub genes potentially associated with rice response to the three Xoo strains. The expression of several hub genes was validated to be induced by all three Xoo strains, suggesting its role in bacterial blight disease response to Xoo strains. Genomic analysis of the Xoo strains belonging to pathotypes IV, V, and IX, identification of effectors and genes related to Xoo virulence in rice plants will provide insights into understanding the molecular mechanism underlying rice-Xoo interaction and the gene expression pattern in response to Xoo infection.

A novel highly virulent nephropathogenic QX-like infectious bronchitis virus originating from recombination of GI-13 and GI-19 genotype strains in China

Infectious bronchitis virus (IBV) is one of the most widely spread RNA viruses, causing respiratory, renal, and intestinal damage, as well as decreased reproductive performance in hens, leading to significant economic losses in the poultry industry. In this study, a new IBV strain designated as CK/CH/GX/LA/071423 was successfully isolated from the 60-day-old Three-Yellow chicken vaccinated with H120 and QXL87 vaccines. The complete genome sequence analysis revealed that the CK/CH/GX/LA/071423 strain shared a high similarity of 96.7% with the YX10 strain belonging to the GI-19 genotype. Genetic evolution analysis based on the IBV S1 gene showed that the CK/CH/GX/LA/071423 isolate belonged to the GI-19 genotype. Recombination analysis of the virus genome using RDP and Simplot software indicated that CK/CH/GX/LA/071423 was derived from recombination events between the YX10 and 4/91 vaccine strains, which was supported by phylogenetic analysis using gene sequences from the 3 regions. Furthermore, the S1 protein tertiary structure differences were observed between the CK/CH/GX/LA/071423 and the QXL87 and H120 vaccine strains. Pathogenicity studies revealed that the CK/CH/GX/LA/071423 caused death and led to pale and enlarged kidneys with abundant urate deposits, indicative of a nephropathogenic IBV strain. High virus titers were detected in the trachea, kidneys, and cecal tonsils, demonstrating broad tissue tropism. Throughout the experimental period, the virus positive rate in throat swabs of the infected group reached to 100%. These findings highlight the continued predominance of the QX genotype IBV in Guangxi of China and the ongoing evolution of different genotypes through genetic recombination, raising concerns about the efficacy of current IBV vaccines in providing effective protection to poultry.

The complete chloroplast genome sequence and phylogenetic relationship analysis of Eomecon chionantha, one species unique to China.

Eomecon chionantha Hance, an endemic species in China, has a long medical history in Chinese ethnic minority medicine and is known for its anti-inflammatory and analgesic effects. However, studies of E. chionantha are lacking. In this study, we investigated the characteristics of the E. chionantha chloroplast genome and determined the taxonomic position of E. chionantha in Papaveraceae via phylogenetic analysis. In addition, we determined molecular markers to identify E. chionantha at the molecular level by comparing the chloroplast genomes of E. chionantha and its closely related species. The complete chloroplast genomic information indicated that E. chionantha chloroplast DNA (178,808bp) contains 99 protein-coding genes, 8 rRNAs, and 37 tRNAs. Meanwhile, we were able to identify a total of 54 simple sequence repeats through our analysis. Our findings from the phylogenetic analysis suggest that E. chionantha shares a close relationship with four distinct species, namely Macleaya microcarpa, Coreanomecon hylomeconoides, Hylomecon japonica, and Chelidonium majus. Additionally, using the Kimura two-parameter model, we successfully identified five hypervariable regions (ycf4-cemA, ycf3-trnS-GGA, trnC-GCA-petN, rpl32-trnL-UAG, and psbI-trnS-UGA). To the best of our knowledge, this is the first report of the complete chloroplast genome of E. chionantha, providing a scientific reference for further understanding of E. chionantha from the perspective of the chloroplast genome and establishing a solid foundation for the future identification, taxonomic determination and evolutionary analysis of this species.

Biocontrol performance of a novel Bacillus velezensis L33a on tomato gray mold and its complete genome sequence analysis

As one of the most destructive diseases, gray mold caused by Botrytis cinerea is harmful to many important economically crops including tomatoes. The use of microorganisms for biological control is considered safe and effective. Here, a newly isolated strain L33a, which showed a 75.22% inhibition ratio against B. cinerea and broad-spectrum resistance to the tested eight phytopathogenic fungi, was characterized by a series of experiments. Strain L33a was identified as Bacillus velezensis based on its biochemical, morphological, and whole-genome profiles. Microstructural analysis revealed that L33a significantly inhibited the mycelial growth of B. cinerea by compromising the integrity of the cell membrane. The acupuncture inoculation assay demonstrated that L33a significantly alleviated disease symptoms caused by gray mold in tomato fruit through downregulation of pathogenicity-related genes of B. cinerea and activation of the transcription of genes involved in the salicylic acid (SA), jasmonic acid (JA), and antioxidant pathways of the tomato fruit. Whole-genome sequencing indicated that the total size of the L33a genome was 4032063 bp, which contained 14 biosynthetic gene clusters reported to be responsible for the biosynthesis of antimicrobial substances. Forty-one compounds were identified in the volatile organic compounds of strain L33a by using Gas Chromatography-Mass Spectrometer (GC-MS) analysis. Among these, 2,3-butanediol, acetoin, 2-tridecanol, and 2-tridecanone were the dominant compounds. Our study revealed that L33a has substantial potential as an effective biocontrol agent, offering valuable insights into postharvest fruit biocontrol.

Effect of volatile compounds produced by Weissella cibaria BWL4 on Botrytis cinerea infection in fruit and complete genome sequence analysis of BWL4

Botrytis cinerea, one of the most harmful pathogenic fungi, causes significant postharvest decay in fruit. In this study, the effects of Weissella cibaria BWL4 on B. cinerea and the gray mold in grape and kiwifruit were evaluated. Additionally, the genome of an antagonistic bacterium BWL4 was sequenced. Results showed that the application of BWL4 inhibited B. cinerea infection in grape and kiwifruit. BWL4 produced antifungal volatile substances against B. cinerea. A gas chromatography–mass spectrometry of solid phase microextraction samples (SPME-GC-MS) analysis showed that BWL4 produced 54 volatiles. Among these volatiles, furfural, 2,6-dimethyl-pyrazine, dimethyl trisulfide, 2-pentyl furan, 2-ethyl hexanol, hexamethyl-cyclotrisiloxane, dodecamethyl-cyclohexasiloxane, 2,4-di-tert-butylphenol, and butylated hydroxytoluene, exhibited antifungal activity against B. cinerea, with MICs of 0.8 g L−1, 0.2 g L−1, 0.9 g L−1, 5 g L−1, 0.4 g L−1, 8 g L−1, 8 g L−1, 80 mg L−1, and 2 g L−1, respectively. In vivo assay showed that these compounds inhibited the disease development in kiwifruit. These compounds except dimethyl trisulfide can also inhibit gray mold in grape. The genome of W. cibaria BWL4 consists of a circular chromosome of 2.42 Mb and 6 plasmids. The genome size is 2.53 Mb, encoding 2327 putative genes. Several genes involved in the biosynthesis of some antifungal volatiles and 17 genes associated with biocontrol were identified. These results suggested that W. cibaria BWL4 can be considered as an effective biological control agent for the postharvest management of B. cinerea infection.

Complete Genome Sequence Analysis of Bacillus subtilis MC4-2 Strain That against Tobacco Black Shank Disease.

The MC4-2 bacterium strain was isolated and purified from the Periplaneta americana intestine as a biocontrol agent with good antagonistic effect against the pathogens of a soil-borne disease called tobacco black shank. The MC4-2 strain was found to have good broad-spectrum inhibition by plate stand-off test. Based on 16S rRNA and gyrB genes, ANI analysis, and other comparative genomics methods, it was determined that the MC4-2 strain was Bacillus subtilis. The complete genome sequence showed that the genome size was 4,076,630 bp, the average GC content was 43.78%, and the total number of CDSs was 4,207. Genomic prediction analysis revealed that a total of 145 genes were annotated by the CAZy, containing mainly GH and CE enzymes that break down carbohydrates such as glucose, chitin, starch, and alginate, and a large number of enzymes involved in glycosylation were present. A total of ten secondary metabolite clusters were predicted, six clusters of which were annotated as surfactin, bacillaene, fengycin, bacillibactin, subtilosin A, and bacilysin. The present investigation found the biological control mechanism of B. subtilis MC4-2, which provides a strong theoretical basis for the best use of this strain in biological control methods and provides a reference for the subsequent development of agents of this bacterium.

Recent Molecular Characterization of Porcine Rotaviruses Detected in China and Their Phylogenetic Relationships with Human Rotaviruses.

Porcine rotavirus A (PoRVA) is an enteric pathogen capable of causing severe diarrhea in suckling piglets. Investigating the prevalence and molecular characteristics of PoRVA in the world, including China, is of significance for disease prevention. In 2022, a total of 25,768 samples were collected from 230 farms across China, undergoing porcine RVA positivity testing. The results showed that 86.52% of the pig farms tested positive for porcine RVA, with an overall positive rate of 51.15%. Through the genetic evolution analysis of VP7, VP4 and VP6 genes, it was revealed that G9 is the predominant genotype within the VP7 segment, constituting 56.55%. VP4 genotypes were identified as P[13] (42.22%), P[23] (25.56%) and P[7] (22.22%). VP6 exhibited only two genotypes, namely I5 (88.81%) and I1 (11.19%). The prevailing genotype combination for RVA was determined as G9P[23]I5. Additionally, some RVA strains demonstrated significant homology between VP7, VP4 and VP6 genes and human RV strains, indicating the potential for human RV infection in pigs. Based on complete genome sequencing analysis, a special PoRVA strain, CHN/SD/LYXH2/2022/G4P[6]I1, had high homology with human RV strains, revealing genetic reassortment between human and porcine RV strains in vivo. Our data indicate the high prevalence, major genotypes, and cross-species transmission of porcine RVA in China. Therefore, the continuous monitoring of porcine RVA prevalence is essential, providing valuable insights for virus prevention and control, and supporting the development of candidate vaccines against porcine RVA.

Identification and Genome Characterization of a Novel Muscovy Duck-Origin Goose Parvovirus with Three Recombinant Regions between Muscovy Duck Parvovirus and Goose Parvovirus

Muscovy duck-origin goose parvovirus (MDGPV) is a new virus resulting from the natural recombination of Muscovy duck parvovirus (MDPV) and goose parvovirus (GPV). Previously identified MDGPV strains were found to have two recombination regions, one in the P9 promoter to the NS region and one in the VP3 gene, or only one recombination in the VP3 gene. In 2022, a novel strain of MDGPV known as 2022FZ was identified from China’s mainland. Complete genome sequence analysis showed that there were three recombination regions in this strain: one located in the P9 promoter-NS (425–612 nt) region, one in the NS2 (1,483–1,824 nt) region, and one in the VP3 (3,124–4,248 nt) region, respectively. The recombination regions in the P9 promoter-NS, NS2, and VP3 genes were substituted with the relevant GPVs sequences, whereas the MDPV virulent strain served as the skeleton in this instance. In addition, the 2022FZ strain had multiple unique aa mutations in the NS protein and the VP protein. The Muscovy duckling challenge test showed that MDGPV-2022FZ is less pathogenic to Muscovy ducklings than two recombinant or sole recombinant MDGPV strains. For the first time, our study identified a three-region recombinant MDGPV strain and detected the novel recombination event in the NS2 gene. These results contribute to our understanding of the pathogenicity and genetic diversity of duck parvovirus.

The first complete genome sequence and genetic evolution analysis of bovine norovirus in Xinjiang, China

Abstract Introduction Viruses are among the main pathogens causing diarrhoea in calves. The current study found that bovine norovirus (BNoV) is one of the principal viruses causing diarrhoea in calves in Xinjiang, China. Material and Methods A total of 974 calf faecal samples from six regions in Xinjiang were tested for BNoV using reverse-transcriptase PCR. The genomic characteristics of BNoV and the genetic evolution of the VP1 gene, protein three-dimensional structure characteristics and amino acid variation were analysed using bioinformatics methods. Results Epidemiological survey results showed that the infection rate of BNoV was 19.82%, and all samples tested positive in five regions. The results of the genetic evolution analysis showed that BNoV strains from Tacheng of northern Xinjiang and Kashgar of southern Xinjiang both belonged to the GIII.2 genotype of BNoV but were not on the same cluster of evolutionary branches. Additionally, the amino acid variation of the VP1 protein was not observed to significantly affect its spatial structure. Conclusion This study is the first to report the genetic characteristics of the BNoV complete genome sequence in Xinjiang and provides a scientific basis for BNoV vaccine development and pathogenesis research.