Effects of gas condition on acetic acid fermentation by Clostridium thermocellum and Moorella thermoacetica (C. thermoaceticum).

Fermentation with acetogens can be affected by cultivation gas phase, but to date, there is not enough evidence on that matter for Clostridium thermocellum and Moorella thermoacetica. In this work, the effects of sparged CO2 as well as sparged and non-sparged N2 on these microorganisms were studied using glucose and cellobiose as substrates. It was revealed that sparged CO2 and non-sparged N2 supported growth and acetic acid production by C. thermocellum and M. thermoacetica, while sparged N2 inhibited both of the microorganisms. Notably, part of the sparged CO2 was fermented by the co-culture system and contributed to an overestimation of the products from the actual substrate as well as an erring material balance. The best condition for the co-culture was concluded to be N2 without sparging. These results demonstrate the importance of cultivation conditions for efficient fermentation by anaerobic clostridia species.

Detection of novel ferret coronaviruses and evidence of recombination among ferret coronaviruses.

In an epidemiological study of ferret coronaviruses (FRCoVs), novel FRCoV strains (Saitama-1 and Aichi-1) were detected by reverse transcription-polymerase chain reaction (RT-PCR) and nucleotide sequence analysis of partial RNA-dependent RNA polymerase (RdRp) genes. Phylogenetic analysis indicated that these strains belonged to different clusters from other FRCoV strains. Next, the nucleotide sequence of the 3'-terminal region of Saitama-1 (8271 bases) strain was determined and compared with those of the other FRCoVs, indicating that the Saitama-1 strain differed from the previously reported MSU-1 and MSU-2 strains in the regions encoding spike (S) protein, nucleocapsid, and open reading frame 7b. Furthermore, the results of SimPlot analysis indicated that FRCoV (MSU-2 strain) emerged via a recombination event of S protein between the MSU-1 and Saitama-1 strains. This mechanism is similar to that responsible for the emergence of type II feline coronavirus. This information will be useful for understanding the pathogenesis of FRCoV in ferrets.

High rates of infection with novel enterovirus variants in wild populations of mandrills and other old world monkey species.

UNLABELLED: Enteroviruses (EVs) are a genetically and antigenically diverse group of viruses infecting humans. A mostly distinct set of EV variants have additionally been documented to infect wild apes and several, primarily captive, Old World monkey (OWM) species. To investigate the prevalence and genetic characteristics of EVs infecting OWMs in the wild, fecal samples from mandrills (Mandrillus sphinx) and other species collected in remote regions of southern Cameroon were screened for EV RNA. Remarkably high rates of EV positivity were detected in M. sphinx (100 of 102 screened), Cercocebus torquatus (7/7), and Cercopithecus cephus (2/4), with high viral loads indicative of active infection. Genetic characterization in VP4/VP2 and VP1 regions allowed EV variants to be assigned to simian species H (EV-H) and EV-J (including one or more new types), while seven matched simian EV-B variants, SA5 and EV110 (chimpanzee). Sequences from the remaining 70 formed a new genetic group distinct in VP4/2 and VP1 region from all currently recognized human or simian EV species. Complete genome sequences were obtained from three to determine their species assignment. In common with EV-J and the EV-A A13 isolate, new group sequences were chimeric, being most closely related to EV-A in capsid genes and to EV-B in the nonstructural gene region. Further recombination events created different groupings in 5' and 3' untranslated regions. While clearly a distinct EV group, the hybrid nature of new variants prevented their unambiguous classification as either members of a new species or as divergent members of EV-A using current International Committee on Taxonomy of Viruses (ICTV) assignment criteria. IMPORTANCE: This study is the first large-scale investigation of the frequency of infection and diversity of enteroviruses (EVs) infecting monkeys (primarily mandrills) in the wild. Our findings demonstrate extremely high frequencies of active infection (95%) among mandrills and other Old World monkey species inhabiting remote regions of Cameroon without human contact. EV variants detected were distinct from those infecting human populations, comprising members of enterovirus species B, J, and H and a large novel group of viruses most closely related to species A in the P1 region. The viral sequences obtained contribute substantially to our growing understanding of the genetic diversity of EVs and the existence of interspecies chimerism that characterizes the novel variants in the current study, as well as in previously characterized species A and J viruses infecting monkeys. The latter findings will contribute to future development of consensus criteria for species assignments in enteroviruses and other picornavirus genera.

Co-circulation of enteroviruses between apes and humans.

A total of 139 stool samples from wild chimpanzees, gorillas and bonobos in Cameroon and Democratic Republic of Congo (DRC) were screened for enteroviruses (EVs) by reverse transcription PCR. Enterovirus RNA was detected in 10 % of samples, comprising eight from 58 sampled chimpanzees (13.8 %), one from 40 bonobos (2.5 %) and five from 40 gorillas (12.2 %). Three viruses isolated from chimpanzees grouped with human isolate EV-A89 and four (four chimpanzees, one gorilla) represented a newly identified type, EV-A119. These species A virus types overlapped with those circulating in human populations in the same area. The remaining six strains comprised a new species D type, EV-D120, infecting one chimpanzee and four gorillas, and a single EV variant infecting a bonobo that was remarkably divergent from other EVs and potentially constitutes a new enterovirus species. The study demonstrates both the circulation of genetically divergent EV variants in apes and monkeys as well as those shared with local human populations.

Functional interaction of the ankylosing spondylitis-associated endoplasmic reticulum aminopeptidase 1 polymorphism and HLA-B27 in vivo.

The association of ERAP1 with ankylosing spondylitis (AS)1 among HLA-B27-positive individuals suggests that ERAP1 polymorphism may affect pathogenesis by altering peptide-dependent features of the HLA-B27 molecule. Comparisons of HLA-B*27:04-bound peptidomes from cells expressing different natural variants of ERAP1 revealed significant differences in the size, length, and amount of many ligands, as well as in HLA-B27 stability. Peptide analyses suggested that the mechanism of ERAP1/HLA-B27 interaction is a variant-dependent alteration in the balance between epitope generation and destruction determined by the susceptibility of N-terminal flanking and P1 residues to trimming. ERAP1 polymorphism associated with AS susceptibility ensured efficient peptide trimming and high HLA-B27 stability. Protective polymorphism resulted in diminished ERAP1 activity, less efficient trimming, suboptimal HLA-B27 peptidomes, and decreased molecular stability. This study demonstrates that natural ERAP1 polymorphism affects HLA-B27 antigen presentation and stability in vivo and proposes a mechanism for the interaction between these molecules in AS.

Potential effects of a human milk oligosaccharide 6'-sialyllactose on angiotensin II-induced aortic aneurysm via p90RSK/TGF-ß/SMAD2 signaling pathway.

The aberrant phenotypic transformation of vascular smooth muscle cells (VSMCs) is a key factor in the formation of aortic aneurysm (AA). This study aimed to explore the effects of 6'-sialyllactose (6'-SL), a human milk oligosaccharide, on angiotensin II (Ang II)-induced VSMC dysfunction and AA formation both in vitro and in vivo. An AA model was established in male C57BL/6 mice challenged with Ang II via osmotic pumps and a lysyl oxidase inhibitor, ß-aminopropionitrile (BAPN), in drinking water. The mice were administered with 6'-SL, FMK (a p90RSK inhibitor), or losartan (as a positive control). In vitro, VSMCs were pretreated with 6'-SL before Ang II stimulation. We found that p90RSK inhibition abolished Ang II/BAPN-induced thoracic AA and abdominal AA formation. Treatment with 100 mg/kg 6'-SL significantly attenuated Ang II/BAPN-induced aortic dilatation. 6'-SL attenuated Ang II-induced collagen deposition, calcification, and immune cell accumulation. Consistently, 6'-SL downregulated p-p90RSK, p90RSK, and p-SMAD2, and mitigated VSMC contractility loss, as indicated by α-SMA expression in vivo. Interestingly, Ang II-induced transforming growth factor-beta (TGF-ß) signaling pathway was suppressed by p90RSK inhibition in VSMCs. 6'-SL treatment significantly reduced TGF-ß/SMAD2 targets, including dedifferentiation markers such as osteopontin and vimentin, and elastin degradation factors MMP2 and MMP9. Overexpression of p90RSK in VSMCs enhanced TGF-ß and abrogated the effects of 6'-SL. Furthermore, 6'-SL co-treatment abolished high phosphate-induced calcification in vitro via p90RSK/TGF-ß signaling pathway. Altogether, our findings suggest that 6'-SL could be a potential therapeutic candidate for protecting against Ang II-induced AA formation by inhibiting the p90RSK/TGF-ß/SMAD2 signaling pathway.

Ginsenoside Rh1 Inhibits Angiotensin II-Induced Vascular Smooth Muscle Cell Migration and Proliferation through Suppression of the ROS-Mediated ERK1/2/p90RSK/KLF4 Signaling Pathway.

Vascular smooth muscle cell (VSMC) proliferation and migration play key roles in the progression of atherosclerosis and restenosis. A variety of ginsenosides exert various cardiovascular benefits. However, whether and how ginsenoside Rh1 (Rh1) inhibits VSMC dysfunction remain unclear. Here, we investigated the inhibitory effects of Rh1 on rat aortic smooth muscle cell (RASMC) migration and proliferation induced by angiotensin II (Ang II) and the underlying mechanisms. Cell proliferation and migration were evaluated using sulforhodamine B and wound-healing assay. The molecular mechanisms were investigated using Western blotting, quantitative reverse-transcription polymerase chain reaction analysis, immunofluorescence staining, and luciferase assay. Reactive oxygen species (ROS) production was measured using dihydroethidium and MitoSOX staining. We found that Rh1 dose-dependently suppressed Ang II-induced cell proliferation and migration. Concomitantly, Ang II increased protein levels of osteopontin, vimentin, MMP2, MMP9, PCNA, and cyclin D1, while these were reduced by Rh1 pretreatment. Notably, Ang II enhanced both the protein expression and promoter activity of KLF4, a key regulator of phenotypic switching, whereas pretreatment with Rh1 reversed these effects. Mechanistically, the effects of Rh1 on VSMC proliferation and migration were found to be associated with inhibition of ERK1/2/p90RSK signaling. Furthermore, the inhibitory effects of Rh1 were accompanied by inhibition of ROS production. In conclusion, Rh1 inhibited the Ang II-induced migration and proliferation of RASMCs by suppressing the ROS-mediated ERK1/2/p90RSK signaling pathway.

6'-Sialylactose abolished lipopolysaccharide-induced inflammation and hyper-permeability in endothelial cells.

Disruption of the endothelial barrier function and reduction in cell migration leads to endothelial dysfunction. One of the most abundant human milk oligosaccharides, 6'-sialylactose (6'-SL), is reported to exert various biological functions related to inflammatory responses. In this study, we evaluated the effects of 6'-SL on lipopolysaccharide (LPS)-induced inflammation caused by endothelial barrier damage. Our results showed that LPS at 500 ng/mL strongly not only abolished cell migration but also hyperactivated MAPK and NF-κB pathways. 6'-SL suppressed LPS-induced endothelial inflammation via ERK1/2, p38, and JNK MAPK pathways. 6'-SL supported endothelial junctions by upregulating PECAM-1 expression and mRNA levels of tight junctions, such as ZO-1 and occludin, which were downregulated by LPS stimulation. It significantly inhibited the nuclear translocation of NF-κB, along with the downregulation of inflammatory cytokines, including TNF-α, IL-1ß, MCP-1, VCAM-1, and ICAM-1. Furthermore, 6'-SL abolished NF-κB-mediated STAT3 in controlling endothelial migration and hyperpermeability via downregulating STAT3 activation and nuclear translocation. Finally, LPS induced over-expression of VCAM-1 and ZO-1 disassembly in both atheroprone and atheroprotective areas of mouse aorta, which were reversed by 6'-SL treatment. Altogether, our findings suggest that 6'-SL is a potent therapeutic agent for modulating inflammatory responses and endothelial hyperpermeability.

Detection and Characterization of Homologues of Human Hepatitis Viruses and Pegiviruses in Rodents and Bats in Vietnam.

Rodents and bats are now widely recognised as important sources of zoonotic virus infections in other mammals, including humans. Numerous surveys have expanded our knowledge of diverse viruses in a range of rodent and bat species, including their origins, evolution, and range of hosts. In this study of pegivirus and human hepatitis-related viruses, liver and serum samples from Vietnamese rodents and bats were examined by PCR and sequencing. Nucleic acids homologous to human hepatitis B, C, E viruses were detected in liver samples of 2 (1.3%) of 157 bats, 38 (8.1%), and 14 (3%) of 470 rodents, respectively. Hepacivirus-like viruses were frequently detected (42.7%) in the bamboo rat, Rhizomys pruinosus, while pegivirus RNA was only evident in 2 (0.3%) of 638 rodent serum samples. Complete or near-complete genome sequences of HBV, HEV and pegivirus homologues closely resembled those previously reported from rodents and bats. However, complete coding region sequences of the rodent hepacivirus-like viruses substantially diverged from all of the currently classified variants and potentially represent a new species in the Hepacivirus genus. Of the viruses identified, their routes of transmission and potential to establish zoonoses remain to be determined.

Characterization of canine coronavirus spread among domestic dogs in Vietnam.

Canine coronavirus (CCoV) is an important pathogen that causes enteritis in dogs, but there is no information on CCoV infection in Vietnam. To examine the prevalence of CCoV infection among Vietnamese dogs, 201 serum samples were analyzed by virus-neutralization (VN) test. The results showed that antibody against CCoV-II was present in 87 dogs (43.3%). To detect genes of CCoV, fecal samples collected from 30 diarrheic and 50 healthy dogs were examinated by RT-PCR, confirming that 2 diarrheic dogs and 5 healthy dogs were positive for CCoV. Nucleotide sequences of N-terminal region of spike (S) gene indicated that CCoV strains were divided into two subgenotypes, CCoV-IIa and -IIb, respectively. Furthemore, we succeeded in isolating CCoV/dog/HCM47/2015, the isolate was plaque-purified three times, and 3'-terminal one-third of the genome was analyzed. Interestingly, the plaque-purified virus had a large deletion in ORF3abc and E genes (1,165 nt), and a short deletion in ORF7b gene (60 nt), suggesting that these regions are not necessary for in vitro replication of CCoV. Next, the antigenicity between the isolated CCoV-IIb and the other CCoV-IIa was compared by VN test, revealing that antigenicty of the isolated CCoV is equal or higher than that of the other CCoV. In summary, two subgenotypes of CCoV-II are spreading among Vietnamese dogs. The isolated virus with a large deletion after in vitro passage may be useful for the development of vaccine, owing to its antigenicity and efficient viral growth in vitro.

Influenza A virus infection in Japanese wild boars (Sus scrofa leucomystax).

Serum samples were collected from 385 wild boars between 2010 and 2013 to examine the seroprevalence of influenza A virus (IAV) in Japan. Antibodies against IAV were identified using a commercial kit in 13 wild boars (3.4%). To identify the serotypes, positive sera were examined by virus-neutralization test using representative serotypes and strains. Three wild boars in Yamaguchi and four in Tochigi showed the highest antibody titers against the pandemic H1N1 2009 virus and classical swine H1N1 virus strains, respectively. These data indicate that wild boars may have close contact with humans and domestic pigs and therefore that there is potential for IAVs to reassort in wild boars as they have been shown to do in pigs.

Peptides eluted from HLA-B27 of human splenocytes and blood cells reveal a similar but partially different profile compared to in vitro grown cell lines.

The sequences and profiles of peptides which bind to HLA-B*2705 splenocytes and peripheral blood cells were compared with those previously published from in vitro long-term cell cultures. B*2705 peptide profile analysed by solid-phase Edman degradation and 15 individual peptide sequences determined by LC-MS/MS were partially similar to those defined from in vitro long-term cell cultures. Arg at P2 was found in 11 of 15 sequenced peptides (73.3%). This value is lower in comparison with other published data. Two sequences were matching to unknown proteins, which displayed similarity with myosin. These are first data on peptide sequences isolated directly from HLA-B27 molecules without prior in vitro propagation of the cells.