VP1 Gene Research Articles

Two novel sites determine genetic relationships between CPV-2 and FPV: an epidemiological survey of canine and feline parvoviruses in Changchun, China (2020)

Canine parvovirus (CPV-2) and feline parvovirus (FPV) cause severe hemorrhagic diarrhea disease in dogs, cats, and fur-bearing and wildlife carnivores worldwide, continuing to pose significant threats. In this study, 140 rectal swabs were collected from 70 domestic dogs and 70 cats with clinical diarrhea in veterinary clinics in Changchun during 2020. A total of 64.3% (45/70) of dogs and 55.7% (39/70) of cats tested positive for CPV-2 or FPV using colloidal gold strips. Amino acid (aa) sequence alignment of the VP2 protein from 39 CPV-2 and 36 FPV samples revealed that 79.5% (31/39) were CPV-2c, 17.9% (7/39) were a new CPV-2a, and 2.6% (1/39) were mink enteritis virus (MEV). and 8.3% (3/36) FPV from the cats was infected by CPV-2, which suggested that CPV-2c was the dominant variant in dogs and FPV was the major pathogen in cats in Changchun city. Phylogenetic relationships of VP2 genes showed that 26 parvoviruses were closely related to domestic strains previously published in China; however, 8 FPVs and CPV-JL-15/China/2020 were clustered in the lineage of South Asiatic and European countries, and 7 out of 8 FPVs were close to Italy. In addition to Q247H, I248Y, F544Y, and E/V545V/K, two novel site mutations of N23D or L630P in NS1 protein, associated with viral cross-species transmissions, were first found as a reminder of genetic relationships of CPV-2 variants and FPVs in the same branch. Thus, regular and massive virus surveillance of parvovirus is necessary to cope with its ongoing infection, circulation, mutations, and evolutions to new subtypes with strong survival abilities.

New Genotype G3 P[8] of Rotavirus Identified in a Mexican Gastroenteric Rabbit

Rotavirus species A (RVA) is a major cause of acute viral gastroenteritis in young humans and diverse animal species. The study of the genetic characteristics of RVAs that infect rabbits (Oryctolagus cuniculus) (lapine strain [LRV]) has been limited, and, to date, the most common and epidemiologically important combinations of G/P genotypes in rabbits have been reported to be G3 P[14] and G3 P[22]. In this study, a rotavirus species A detected from an outbreak of enteritis in a Mexican commercial rabbitry was genotypically characterized. Based on sequence and phylogenetic analysis of the VP7 and VP4 genes, the strain identified in this study (C-3/15) demonstrated a G3 P[8] genotype of rotavirus, which had not previously been reported in rabbits. Moreover, both genes were closely related to human, not lapine, rotaviruses. The G3 genotype has been reported in a wide variety of hosts, including humans and rabbits, whereas the P[8] genotype has only been reported in humans. Because this combination of genotypes has never been identified in rabbits, it is proposed that the finding presented here is possibly the result of an interspecies transmission event. This is the first work to study the molecular characteristics of rotaviruses in rabbits in Mexico, as well as the identification of human G3 and P[8] genotypes in a rabbit with enteric disease.

Molecular characterization of VP1 gene during the foot and mouth disease virus outbreak in East Java, Indonesia, in 2022

Molecular characterization of VP1 gene during the foot and mouth disease virus outbreak in East Java, Indonesia, in 2022

Human parvovirus B19 virus-like particle formation in Nicotiana benthamiana

There has been a surge in the interest to utilize plants as hosts for producing vaccine antigens. In this study, we demonstrated the successful expression of the human parvovirus B19 (B19V) capsid protein (VP2) in Nicotiana benthamiana cells. The B19V VP1 and VP2 genes were cloned under the control of estrogen-inducible promoters and transiently expressed in N. benthamiana leaves using the agroinfiltration method. The addition of estrogen significantly boosted the expression of VP2. Furthermore, codon optimization of the VP2 sequence resulted in over a 30-fold increase in its expression compared with that of the wild-type. Analysis of negatively stained samples by sucrose density gradient ultracentrifugation and electron microscopy revealed that the expressed VP2 proteins formed spherical particles with diameters of approximately 20 nm. Immunostaining analysis of protoplasts derived from VP2-expressing N. benthamiana leaves indicated that VP2 signals were predominantly localized in the cytoplasm. These findings strongly suggested that B19V VP2 assembles and formed virus-like particles (VLPs) within the cytoplasm of N. benthamiana cells, presenting a promising method for producing B19V VLPs in plant systems.

Genetic Diversity and Recombination Analysis of Canine Parvoviruses Prevalent in Central and Eastern China, from 2020 to 2023

Canine parvovirus type-2 (CPV-2), the primary causative agent of serious canine enteric diseases, is highly contagious and associated with high fatality rates worldwide. To comprehend the current emergence of CPV-2 in central and eastern China, 130 rectal swabs from domestic or stray dogs with gastroenteritis symptoms were collected during 2020–2023. A total of 118 positive samples were detected via polymerase chain reaction, and further used to amplify and sequence the VP2 gene. Sequence analysis of the deduced amino acids of VP2 protein indicated that CPV-2c was the most prevalent variant (n = 106, 89.83%), followed by the novel CPV-2a (n = 10, 8.47%) and CPV-2b (n = 2, 1.69%) variants. The VP2 protein from the obtained and reference strains showed 86.95% (AH2103 and HB2108) to 99.94% identity. Based on the nine predicted recombination events, some prevalent CPV-2c strains were highly similar to previously isolated strains, indicating their complex evolution and recombination. The predicted analysis suggested that mutations in the antigen epitope (Val219Ile, Phe267Tyr, and Asn426Glu) and other mutations (Met87Leu, Ile101Thr, and Ser297Ala) affect the tertiary structure of the VP2 protein. This research will help us understand the recent evolution and mutation of Chinese CPV-2 and provide suggestions for updating the CPV-2 vaccine.

Detection and molecular characterization of avian polyomavirus in budgerigar and non-budgerigar psittacine species in bird markets of Pakistan

Avian Polyomaviruses are imposing severe health problems in budgerigars, non-budgerigar Psittacine species, and non-psittacine species all over the world, including Pakistan. It marks future challenges for aviculturists and pet store owners, causing significant financial losses. This study emphasizes the occurrence and molecular characterization of polyomaviruses in budgerigars and non-budgerigar Psittacine species. Thirty-five feather Samples of adult birds and 15 tissue samples of deceased birds were collected for the detection of Avian polyomavirus based on the VP1 gene. Screening of samples by PCR revealed the presence of 550 bp VP1 gene in deceased nestlings of two lovebirds and four budgerigars, while the feather samples of adult birds were all negative for VP1 gene. The overall positive rate of APV in Psittacine birds was 6/50 (12 %), and the distribution frequency of virus among species was 4/19 (20 %) in Budgerigars and 2/31 (6.4 %) in non-budgerigar. Positive samples were subjected to partial sequencing which showed a nucleotide similarity index of VP1 gene between 97.46 % & 99.6 % with reference sequences in GenBank. The main problem that researchers are dealing with is the scarcity of data on the prevalence and identification of APV in Pakistan. This study is a milestone for further research on APV for the diagnosis and development of vaccines.

McHDV VP60 Virus-like Particles Elicit Protective Immunity Against Moschus chrysogaster Hemorrhagic Disease in Rabbits

Moschus chrysogaster viral hemorrhagic disease (McVHD), caused by the Moschus chrysogaster hemorrhagic disease virus (McHDV), is an acute and highly fatal infectious disease of musk deer. At present, there is no prevention or treatment for this disease. In this study, we constructed a recombinant bacmid containing the McHDV VP60 gene and obtained the recombinant baculovirus rBac-McHDV VP60 by transfection into Sf9 (Spodoptera frugiperda) insect cells. The McHDV VP60 protein was successfully expressed in the insect cell-baculovirus expression system; furthermore, it was released in the supernatant of infected insect cells and spontaneously assembled to form virus-like particles (VLPs), which were structurally and immunologically indistinguishable from the Moschus chrysogaster viral hemorrhagic disease virion. Hypodermic vaccination of rabbits with the VLPs conferred complete protection in 14 days; this protection was found to be effective from the seventh day after VLP injection and was accompanied by a strong humoral response. This study is the first attempt to express the VP60 gene of McHDV using an insect baculovirus system, which provides an experimental basis for the virus-like particle vaccine of McVHD.

Development of a Quadruplex RT-qPCR for the Detection of Feline Kobuvirus, Feline Astrovirus, Feline Bufavirus, and Feline Rotavirus

Feline kobuvirus (FeKoV), feline astrovirus (FeAstV), feline bufavirus (FeBuV), and feline rotavirus (FRV) are important pathogens for gastroenteritis, which is characterized by vomiting, diarrhea, and dehydration. Four pairs of primers and probes were designed to target the FeKoV VP1, FeAstV ORF2, FeBuV VP2, and FRV NSP4 genes, and a quadruplex real-time quantitative RT-PCR (RT-qPCR) assay capable of the simultaneous detection of four feline enteroviruses was developed after optimization of reaction conditions. The established quadruplex RT-qPCR assay showed high specificity, sensitivity, and reproducibility. The assay could detect and discriminate FeKoV, FeAstV, FeBuV, and FRV, but not other feline-related pathogens. The limits of detection (LODs) of FeKoV, FeAstV, FeBuV, and FRV were 109.761, 115.834, 125.481, and 113.875 copies/reaction, respectively. The intra- and inter-assay coefficients of variation (CV) were 0.15–1.61% and 0.15–1.59%, respectively. In all, 1869 clinical samples from Guangxi province in Southern China were tested using the developed assay, and the positivity rates of FeKoV, FeAstV, FeBuV, and FRV were 1.93%, 9.36%, 0.32%, and 0.75%, respectively. These samples were also tested using reference assays, and the coincidence rates of the results between the developed and reference methods were 99.63% (FeKoV), 98.72% (FeAstV), 100% (FeBuV), and 100% (FRV), respectively. The results indicated that the developed assay could provide a new detection method for these four viruses associated with feline gastroenteritis.

Development and application of a quadruplex real-time PCR method for Torque teno sus virus 1, Porcine circovirus type 2, pseudorabies virus, and porcine parvovirus.

In clinical diagnosis of porcine diseases, co-infection with multiple viruses often leads to similar clinical symptoms. Postweaning multisystemic wasting syndrome (PMWS) can be caused by infections with TTSuV or PCV2, while PCV2, PRV, and PPV can cause respiratory and reproductive disorders in pigs. The overlapping clinical and pathological features of these infections necessitate the development of a rapid and specific method for differentiating and detecting these four DNA viruses. In this study, four pairs of primers and TaqMan probes were designed targeting the conserved sequence of TTSuV, the Rep gene of PCV2, the gE gene of PRV, and the VP2 gene of PPV. After optimizing reaction conditions, including annealing temperature, primer concentration, and probe concentration, a quadruplex real-time PCR method was developed. This method can specifically detect TTSuV1, PCV2, PRV, and PPV simultaneously, with no cross-reactivity with ASFV, CSFV, PRRSV, PEDV, PSV, and TGEV. The minimum detection limit for each virus was 10 copies/μl, and the inter-assay and intra-assay coefficients of variation ranged from 0.33% to 1.43%. Subsequently, 150 clinical samples were tested to evaluate the practical applicability of this method. The positive rates for TTSuV1, PCV2, PRV, and PPV were 8.6% (13/150), 10.67% (16/150), 14% (21/150), and 11.33% (17/150), respectively. The results indicate that the established quadruplex real-time PCR method can assist in the accurate and rapid diagnosis of TTSuV1, PCV2, PRV, and PPV in clinical settings, providing robust support for the prevention and control of these infections.

Comparative pathogenicity of duck hepatitis A virus genotype 3 in different duck breeds: Implications of the diagnosis and prevention of duck viral hepatitis

Duck hepatitis A virus (DHAV) infection in ducklings causes acute hepatitis with considerable economic losses. In this study, Pekin and Muscovy duckling flocks (n=9) suffering from high mortality and hepatic lesions were examined by RT-PCR for DVHA. 44.4 % (4/9) of samples were positive for DHAV (5′ UTR region), of which 100 % (4/4) were DVHA-3 (VP1 gene). VP1 sequencing and phylogenetic analysis of an isolate originated from Muscovy ducklings showed that it shared 96.8 % −100 %, 88.5–89.2 %, and 86.5–88.2 % nucleotide similarity (ns) with the Egyptian, Korean-Vietnamese, and Chinese DVHA-3 strains, respectively. It was distinguished from the DHAV-1 vaccine (67.6 % ns). The sequenced DVHA-3 isolate was used to experimentally infect 5-day-old Pekin and Muscovy ducklings vs. control groups. No apparent clinical signs or deaths were reported in the experimentally-infected groups. Pekin ducklings showed greater cloacal viral shedding than Muscovy until the 6th dpi (P<0.05). DVHA-3 induced a significant rise in IFN-β and IL-1β serum levels, where Muscovy ducklings' levels were higher than Pekin ducklings. Among the biochemical parameters, AST was only increased on the 4th dpi in both breeds vs. control (P<0.05). Compared to Muscovy ducklings at 2, 4, and 6 day post infection (dpi), the infected Pekin group had lower lipase levels (P≥0.05, p<0.05, and p<0.05, respectively), while ALT was higher at 4 and 6 dpi (P<0.05). The histopathological lesions supported the gross lesions, and their scores were dominant at 2 and 4 dpi in both breeds. At 6 and 8 dpi, Pekin showed more severe histopathological changes compared to Muscovy for the liver, heart, brain, and intestines; the pancreas, kidney, and lung showed the opposite pattern. In conclusion, Pekin ducklings displayed distinct DHAV-3 infection results from Muscovy ducklings, and more research utilizing a variety of DHAV-3 strains has to be carried out.

Immunogenicity analysis based on VP1 and VP2 proteins of bovine enterovirus

Bovine enterovirus (BEV) infection manifests as a spectrum of clinical signs affecting the respiratory, gastrointestinal, and reproductive systems in cattle. Outbreaks of this disease results in large economic losses to the bovine industry worldwide. Currently there are no efficacious vaccines and medicines to prevent BEV infection. In the present study, reverse transcription-polymerase chain reaction was used to amplify the VP1 and VP2 genes of BEV, enabling the synthesis of a chimeric recombinant protein which contained partial sequences from both proteins. Subsequently, the emulsified purified proteins with Freund’s adjuvant were used for triple-fold immunization of 4-week-old Institute of Cancer Research (ICR) mice. The mice were subsequently subjected to a challenge assay which elicited an immune response that was characterized by elevated titers of BEV-specific neutralizing antibodies. Notably, the results showed that the purification of pET32a-VP1 and pET32a-VP2 proteins markedly curtailed virus excretion and mitigated the histopathological damage usually associated with BEV infections. These were observed in the small intestines, kidneys and brain in infected animals. It also alleviated clinical symptoms such as hypothermia and weight loss. In summary, this study offers a theoretical and practical basis for BEV vaccine development.

Molecular typing of Protoparvovirus carnivoran 1 in Egyptian cats diagnosed with feline panleukopenia

Feline panleukopenia (FPL) in cats is caused by either feline parvovirus (FPV) or canine parvovirus (CPV-2), which belong to the same species “Protoparvovirus carnivoran 1”. While FPV is widely recognized as the principal cause of FPL, CPV-2 has been detected at a higher rate than FPV in sick cats in a recent Egyptian study. To assess this conflict, the present study aimed to determine which Protoparvovirus carnivoran 1 is commonly associated with FPL in Egyptian cats. From Dec-2022 to Jan-2024, 43 cats presenting with acute gastroenteritis and testing positive for FPL using in-clinic assay, SNAP® parvo, were tested for Protoparvovirus carnivoran 1 DNA using conventional PCR. Typing of Protoparvovirus carnivoran 1 was conducted by partial VP2 gene sequencing. Additional epidemiological aspects of the disease were investigated, including seasonal pattern, case-fatality rate, median survival time to death, and the association between FPL outcomes and selected factors like age, sex, vaccination status, and clinical signs (vomiting and diarrhea). All cats tested positive for Protoparvovirus carnivoran 1 DNA and FPV was detected in all cats with strong PCR amplicons (n=39). The following seasonal pattern was recorded: cases emerging in autumn, peaking during winter, declining in spring, and disappearing in summer. The case-fatality rate was 41.6%, and the median time to death was two days. None of the studied factors affected FPL outcomes. In conclusion, FPL in Egyptian cat populations is primarily caused by FPV, not CPV-2, and is particularly prevalent in winter.

Molecular Detection and Phylogenetic Tree of Rotavirus from Human and Sheep, Iraq

Group A rotaviruses (RVA) have been linked to acute gastroenteritis in children, as well as young domestic and wild animals. In Iraq few molecular studies involved the genotype profile of rota virus and this is the first study target VP1 gene. Data were collected from January to February 2018 and subsequently examined utilizing a PCR and because of low viral concentration we utilized Nested PCR. Sequenced PCR result at 327 bp fragment in 4 A rota virus-positive samples deposited in GenBank under accession number (MH11809.1, MH118097.1) for human and (MH118095.1, MH118094.1) to sheep. Children samples were identical to the other human A rota virus at (99-100%), with Japan feline-like human G6P and G6P[14] isolated from cattle as well as, animal rota virus (99-100%) similarity with cross relation with human Wa-Like and porcine, moreover Morocco characterized caprine, bovine rotavirus and Slovenian Roe deer ARV.

Whole Genome Sequences of the Wildtype AU-1 Rotavirus A Strain: The Prototype of the AU-1-like Genotype Constellation.

Most human rotaviruses belong to the Wa-like, DS-1-like, or AU-1-like genotype constellation. The AU-1-like constellation, albeit minor, captured attention because its prototype strain AU-1 originated from feline rotavirus, leading to the concept of interspecies transmission of rotavirus. The AU-1 genome sequence determined by various laboratories over the years has documented two conflicting VP7 sequences in the GenBank. As culture-adaptation may introduce changes in the viral genome, the original fecal (wild-type) and the seed stock of culture-adapted AU-1 genomes were sequenced using the Illumina's MiSeq platform to determine the authentic AU-1 sequence and to identify what mutational changes were selected during cell-culture adaptation. The wild-type and culture-adapted AU-1 genomes were identical except for one VP4-P475L substitution. Their VP7 gene was 99.9% identical to the previously reported AU-1 VP7 under accession number AB792641 but only 92.5% to that under accession number D86271. Thus, the wild-type sequences determined in this study (accession numbers OR727616-OR727626) should be used as the reference. The VP4-P475L mutation was more likely incidental than inevitable during cell-culture adaptation. This was the first study in which the whole genomes of both wild-type and cultured RVA strains were simultaneously determined by deep sequencing.

Wide circulation of type 1 vaccine-derived poliovirus strains in clinical specimens from suspected cases of poliomyelitis, their contacts and in wastewater in Madagascar since late 2020

Madagascar has faced three major outbreaks of vaccine-derived polioviruses (VDPVs) in recent decades, with VDPV type 1 reemerging in late 2020. Here, we report the molecular characterization of these cVDPV1 strains. WHO protocols were used for poliovirus detection in stool and wastewater samples. Molecular genotyping was based on the 5′ non-coding (5′NC), VP1, and 3Dpol regions. From 2020 to 2022, 92 of 5690 stool samples and 129 of 1046 wastewater samples tested positive for cVDPV1. Genetic analysis of the VP1 gene revealed 1.3%–6.1% variability compared to the Sabin strain. Most sequences showed mutations at neurovirulence attenuation sites. Phylogenetic analysis distributed strains into four genogroups originating from Southern Madagascar. All analyzed cVDPV1 strains were recombinant, containing mutated oral polio vaccine sequences in VP1 and type C enterovirus sequences in other regions. This study demonstrated that all strains were closely related during this epidemic.

Establishment and Application of Mismatch Amplification Mutation Assay-PCR for Rapid Detection and Differentiation of Duck Hepatitis A Virus-1 Attenuated Vaccine and Wild Strains.

Duck hepatitis A virus type 1 (DHAV-1) is the main pathogen causing viral hepatitis in ducks, marked by high contagion and acute mortality. Live attenuated DHAV-1 vaccines are widely used to control the disease. This study aims to develop a mismatch amplification mutation assay (MAMA)-PCR for the rapid detection and differentiation of Korean DHAV-1 wild-type strains from vaccine strains. A MAMA primer was designed to target a single nucleotide polymorphism (SNPs) at position 2276 within the VP1 gene, allowing differentiation in a single PCR reaction. The MAMA-PCR accurately identified both strains, with detection limits of 100.5 ELD50/mL and 102.3 ELD50/mL, respectively. The MAMA-PCR demonstrated specificity, showing no cross-reactivity with 12 other viral and bacterial pathogens. The MAMA-PCR was applied to 89 farms, yielding results consistent with nested-PCR and sequence determination, identifying four positive farms for DHAV-1 vaccine strains. In conclusion, this study is the first to employ the MAMA-PCR method to distinguish between DHAV-1 wild-type and vaccine strains. The developed method is rapid, simple, specific, and sensitive, thereby serving as an effective tool for clinical diagnostics in identifying and differentiating between Korean DHAV-1 wild-type and vaccine strains.

Full genome sequence analysis of the predominant and uncommon G9P[4] rotavirus strains circulating in Tehran, Iran, 2021–2022: Evidence for inter and intra-genotype recombination

Group A rotaviruses (RVAs) are a major cause of acute gastroenteritis in children under 5 years of age worldwide. Herein, the genetic sequences of 11 RNA segments from three uncommon G9P[4] RVA strains found in the stool samples of children under 5 years of age in Iran were analyzed using next-generation sequencing (NGS) technology. The genomic constellations of these three uncommon G9P[4] strains indicated the presence of the double and quadruple reassortants of two G9P[4] strains, containing the VP7/NSP2 and VP7/VP2/NSP2/NSP4 genes on a DS-1-like genetic background, respectively. The genome of one strain indicated a Wa-like genetic backbone in a single-reassortant with the VP4 of the DS1-like human strains. With the exception of VP1, VP2, VP7, NSP2, NSP3, and NSP4 genes, which clustered with RVA of human origins belonging to cognate gene sequences of genogroup 1/2 genotypes/lineages, the remaining five genes (VP8/VP4, VP3, VP6, NSP1, NSP5) displayed direct evidence of recombination. It is presumed that the presence of uncommon G9P[4] strains in Iran is not linked to vaccination pressure, but rather to the high prevalence of RVA co-infection or the direct import of these uncommon RVA reassortants strains from other countries (especially those that have implemented RV vaccination).

Screening high-efficiency promoter to construct trans-vp28 gene Anabaena sp. PCC7120 against white spot syndrome virus of Litopenaeus vannamei

This study aimed to select high-quality promoters to construct trans-vp28 gene Anabaena sp. PCC7120 and feed Litopenaeus vannamei to assess the effect of L.vannamei against white spot syndrome virus (WSSV). Transgenic algae were created using five plasmids containing PrbcL, Pcpc560, Ptrc, Ptac, and PpsbA. According to the gene expression efficiency and the growth index of transgenic algae, Pcpc560 was determined to be the most efficient promoter. Shrimps were continuously fed trans-vp28 gene Anabaena sp. PCC7120 for one week and then challenged with WSSV. After the challenge, the transgenic algae group (vp28-7120 group) was continuously immunized [continuous immunization for 0 days (vp28-7120-0d); continuous immunization for 2 days (vp28-7120-2d); continuous immunization for 4 days (vp28-7120-4d)]. After seven days, the daily survival rate of each experimental group was continuously tracked. Following the viral challenge, the hepatopancreas samples were assayed for their levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), thioredoxin peroxidase (TPX), acid phosphatase (ACP), and alkaline phosphatase (AKP) at varying time intervals. In comparison to the positive control group (challenge and no vaccination) and the wild-type group (challenge, fed wild-type Anabaena sp. PCC7120), the vp28-7120 group (challenge, fed trans-vp28 gene Anabaena sp. PCC7120) exhibited a remarkable increase in survival rates, reaching 50 % (vp28-7120-0d), 76.67 % (vp28-7120-2d), and 80 % (vp28-7120-4d). Furthermore, the vp28-7120 group consistently displayed significantly higher activities of SOD, CAT, GSH-Px, ACP, and AKP, while exhibiting notably lower TPX activity, when compared to the control group. These results indicate that the Pcpc560 promoter effectively elevated the expression level of the exogenous vp28 gene and spurred the growth of the trans-vp28 gene Anabaena sp. PCC7120. Consequently, trans-vp28 gene Anabaena sp. PCC7120 significantly bolstered the immunity of L.vannamei. Therefore, utilizing the Pcpc560 promoter to develop trans-vp28 gene Anabaena sp. PCC7120 based oral vaccine is highly beneficial for industrial-scale cultivation, advancing its commercialization prospects.

Comparative Analysis of Ligninolytic Potential among Pleurotus ostreatus and Fusarium sp. with a Special Focus on Versatile Peroxidase

Lignocellulosic biomass is contemplated to be an inexpensive and copious feedstock that can be used for numerous industrial applications. However, lignin forms the lignin sheath and provides a physical barrier to enzymatic hydrolysis. In addition, lignin physically blocks cellulase, preventing it from being combined with the substrate in a process known as non-productive binding. Therefore, the depletion of lignin is a crucial method for obtaining fermentable sugars from the lignocellulosic biomass. Different white-rot fungi secrete different sets of lignin-mineralizing enzymes and each fungus secretes one or more of the three enzymes essential for lignin degradation. Among efficient redox enzymes, versatile peroxidase is extensively studied for its ability to degrade aromatics without the need for a mediator or polyvalent catalytic site. However, the presence of versatile peroxidase in F. spp. has not been studied. This study was planned with the objective of screening and comparing the production of versatile peroxidase enzymes from F. spp. and a standard culture of Pleurotus ostreatus MTCC-142. These fungal strains were first screened on solid media containing tannic acid, malachite green, or bromocresol green. The potency index for the tannic acid, malachite green, and bromocresol green on the 16th day of incubation was reported to be 1.28, 1.07, 1.09, and 1.10, respectively. Versatile peroxidase production patterns were investigated under solid state fermentation conditions for a period of 25 days at different temperatures ranging from 10 to 35 °C. The highest versatile peroxidase activity (592 UL−1) in F. sp. was observed at 30 °C after the 7th day of incubation. The molecular confirmation showed the presence of the vp gene in F. sp. along with Pleurotus ostreatus MTCC-142. The results determined that F. sp. possesses a versatile peroxidase enzyme and is able to degrade lignin efficiently, and thus it could be utilized as an alternative to other ligninolytic enzyme-producing fungi.

Genomic surveillance reveals low-level circulation of two subtypes of genogroup C coxsackievirus A10 in Nanchang, Jiangxi Province, China, 2015-2023.

In recent years, coxsackievirus (CV) A10 has been associated with increasing sporadic hand, foot, and mouth disease (HFMD) cases and outbreaks globally. In addition to mild symptoms such as pharyngitis and herpangina, CVA10-related complications or even fatality can occur. Currently, systematic phylogenetic studies of CVA10 are limited. In this study, we first explored the epidemiological and genetic characteristics of CVA10 in Nanchang, an inland southeastern city of China, based on the HFMD surveillance network from 2015-2023. Among 3429 enterovirus-positive cases, 110 (3.04%) were associated with CVA10, with a male-to-female ratio of 1.62. The median age of the CVA10 patients was 2.3 years (interquartile range, IQR 1.0-4.0), with 94.55% (104/110) of the patients aged less than 5 years. Phylogenetic analyses using the full-length VP1, 5'UTR, P1, P2, P3 sequences and near full-length genomes indicated that CVA10 strains (n = 57) isolated in Nanchang belonged to genogroup C; two strains identified in 2017 belonged to C1 subtypes clustered with strains from Vietnam, Madagascar, France and Spain; and the others belonged to C2 subtypes interdigitating with CVA10 isolates from mainland China, the United States and Australia. Through extensive analysis, we identified a rare F168Y mutation in epitope 4 of VP1 in a Madagascar strain of genogroup F and a Chinese strain of genogroup C. Based on Bayesian evolutionary analyses, the average nucleotide substitution rate for the VP1 gene of CV10 strains was 3.07×10-3 substitutions/site/year. The most recent common ancestor (tMRCA) of genogroup C was dated 1990.84, and the tMRCA of CVA10 strains from Nanchang was dated approximately 2003.16, similar to strains circulating in other regions of China, suggesting that the viruses were likely introduced and cryptically circulated in China before the establishment of the HFMD surveillance network. Recombination analysis indicated intertypic recombination of the Nanchang strain with the genogroup G strain in the 3D region. Given the shifting dominance of viral genotypes and frequent recombination events, the existing surveillance system needs to be regulated to enhance genomic surveillance efforts on a more diverse spectrum of genotypes in the future.