Contrasting response of comammox Nitrospira, ammonia oxidising bacteria, and archaea to soil pH and nitrogen inputs.

This study aimed to investigate the effect of soil pH change, and nitrogen amendment on ammonia oxidiser abundance and comammox Nitrospira community composition. The experimental design used soil mesocosms placed in a temperature-controlled incubator for 90 days. A Templeton silt loam was used as its physiochemical properties are typical of the region's dairy farms. The results showed that comammox Nitrospira clade B preferred the natural (pH 6.1-6.2) soil pH with no applied nitrogen. Furthermore, synthetic urine (N700) decreased the abundance of comammox Nitrospira clade B. This may have been because the large amounts of available ammonia in the N700 treatments inhibited the growth of comammox Nitrospira. These results suggest that while comammox Nitrospira clade B are present in New Zealand dairy farm soils, but their role in nitrification in the very high nitrogen environment under a urine patch in grazed pastures may be limited. Further research is needed to confirm this. In contrast to comammox, the AOB community (dominated by Nitrosospira) responded positively to the application of synthetic urine. The response was greatest in the high pH soil (7.1), followed by the natural and then the low pH (4.9) soils. This may be due to the difference in ammonia availability. At high pH, the ammonia/ammonium equilibrium favours ammonia production. Calculated ammonia availability in the N700 treatments accurately predicted the AOB amoA gene abundance. Interestingly, the AOA community abundance (which was predominantly made up of Thaumarchaeota group I.1b clade E) seemed to prefer the natural and high pH soils over the low pH. This may be due to the specific lineage of AOA present. AOA did not respond to the application of nitrogen.

Detection of enterovirus D68 among children with severe acute respiratory infection in Myanmar.

BACKGROUND: Enterovirus D68 (EV-D68) is an important reemerging pathogen that causes severe acute respiratory infection and acute flaccid paralysis, mainly in children. Since 2014, EV-D68 outbreaks have been reported in the United States, Europe, and east Asia; however, no outbreaks have been reported in southeast Asian countries, including Myanmar, during the previous 10 years. METHODS: EV-D68 was detected in nasopharyngeal swabs from children with acute lower respiratory infections in Myanmar. The samples were previously collected from children aged 1 month to 12 years who had been admitted to the Yankin Children Hospital in Yangon, Myanmar, between May 2017 and January 2019. EV-D68 was detected with a newly developed EV-D68-specific real-time PCR assay. The clade was identified by using a phylogenetic tree created with the Bayesian Markov chain Monte Carlo method. RESULTS: During the study period, nasopharyngeal samples were collected from 570 patients. EV-D68 was detected in 42 samples (7.4 %)-11 samples from 2017 to 31 samples from 2018. The phylogenetic tree revealed that all strains belonged to clade B3, which has been the dominant clade worldwide since 2014. We estimate that ancestors of currently circulating genotypes emerged during the period 1980-2004. CONCLUSIONS: To our knowledge, this is the first report of EV-D68 detection in children with acute lower respiratory infections in Yangon, Myanmar, in 2017-2018. Detection and detailed virologic analyses of EV-D68 in southeast Asia is an important aspect of worldwide surveillance and will likely be useful in better understanding the worldwide epidemiologic profile of EV-D68 infection.

Phenotypic and genotypic discrimination of Francisella tularensis ssp. holarctica clades.

Francisella tularensis is the causative agent of tularemia, a zoonotic disease with a wide host range. F. tularensis ssp. holarctica (Fth) is of clinical relevance for European countries, including Germany. Whole genome sequencing methods, including canonical Single Nucleotide Polymorphism (canSNP) typing and whole genome SNP typing, have revealed that European Fth strains belong to a few monophyletic populations. The majority of German Fth isolates belong to two basal phylogenetic clades B.6 (biovar I) and B.12 (biovar II). Strains of B.6 and B.12 seem to differ in their pathogenicity, and it has been shown that strains of biovar II are resistant against erythromycin. In this study, we present data corroborating our previous data demonstrating that basal clade B.12 can be divided into clades B.71 and B.72. By applying phylogenetic whole genome analysis as well as proteome analysis, we could verify that strains of these two clades are distinct from one another. This was confirmed by measuring the intensity of backscatter light on bacteria grown in liquid media. Strains belonging to clades B.6, B.71 or B.72 showed clade-specific backscatter growth curves. Furthermore, we present the whole genome sequence of strain A-1341, as a reference genome of clade B.71, and whole proteomes comparison of Fth strains belonging to clades B.6, B.71 and B.72. Further research is necessary to investigate phenotypes and putative differences in pathogenicity of the investigated different clades of Fth to better understand the relationship between observed phenotypes, pathogenicity and distribution of Fth strains.

Soil moisture is a primary driver of comammox Nitrospira abundance in New Zealand soils.

The objectives of this study were to investigate the abundance and community composition of comammox Nitrospira under: (i) pasture-based dairy farms from different regions, and (ii) different land uses from the same region and soil type. The results clearly showed that comammox Nitrospira were most abundant (3.0 × 106 copies) under the west coast dairy farm conditions, where they were also significantly more abundant than canonical ammonia oxidisers. This was also true in the Canterbury dairy farm. The six land uses investigated were pine monoculture, a long term no input ecological trial, sheep + beef and Dairy, both irrigated and non-irrigated. It was concluded that comammox Nitrospira was most abundant under the irrigated dairy farm (2.7 × 106 copies). Contrary to the current industry opinion, the relatively high abundance of comammox Nitrospira under fertile irrigated dairy land suggests that comammox Nitrospira found in terrestrial ecosystems may be copiotrophic. it was also determined that comammox Nitrospira was more abundant under irrigated land use than their non-irrigated counterparts, suggesting that soil moisture is a key environmental parameter influencing comammox abundance. Comammox abundance was also positively correlated with annual rainfall, further supporting this theory. Phylogenetic analysis of the comammox Nitrospira detected determined that 17 % of the comammox community belonged to a newly distinguished subclade, clade B.2. The remaining 83 % belonged to clade B.1. No sequences from clade A were found.

Comparative analyses of the mitochondrial genomes of the cattle tick Rhipicephalus microplus clades A and B from China.

The cattle tick (Rhipicephalus microplus) is one of the most common ticks parasitizing livestock, causing diseases as the vector of pathogens. In this study, we amplified and sequenced the complete mitochondrial (mt) genome of R. microplus from Hainan province of China and compared it with that of R. microplus from Guizhou province of China. The mt genome sequence of R. microplus from Hainan isolate was 15,163 bp in size, which was significantly longer (299 bp) than R. microplus from Guizhou isolate. Nucleotide sequence difference in the entire mt genome except for non-coding region was 5.6% between R. microplus from Hainan and Guizhou isolates. For the 13 protein-coding genes, this comparison revealed the sequence differences of nucleotide (3.8-10.1%) and amino acid (1.2-17.3%). Phylogenetic analysis of R. microplus indicated that R. microplus from Hainan isolate clustered in clade A, and R. microplus from Guizhou isolate clustered in clade B. Taken together, the findings support the recent proposal the existence of two lineages (clades A and B) of R. microplus in China.

Levels of SERPIN family proteins in peri-implant crevicular fluid in patients with peri-implantitis.

PURPOSE: Serine protease inhibitors (SERPINs) family has been discovered in many disorders with proteolysis mechanisms. Our study determined the SERPINBs protein expression via public-based GEO databases and further validated by peri-implant crevicular fluid (PICF) of peri-implantitis patients and healthy recruiters. METHODS: This study is a retrospective analysis. A total of 123 participants of Fujian Medical University Fujian Stomatological Hospital, consisting of 58 cases of peri-implantitis and 65 samples of healthy control were retrospectively analyzed by ELISA assays and explored the gene enrichment pathways and clinical significance of SERPINBs expression accompanied by two different cytokines (IL-6 and TNF-α). Moreover, the clinical significance of SERPINBs was evaluated in peri-implantitis patients with PICF samples by the receiver operating curve (ROC) using the area under the curve (AUC). RESULTS: KEGG database showed that Starch and sucrose metabolism, Retrograde endocannabinoid signaling, Prion diseases, Pentose phosphate pathways, and Olfactory pathways are up-regulated; GO database showed that synapse organization, synapse assembly, sequestering of triglyceride, sensory perception of smell, and regulation of synapse organization pathways are up-regulated. SERPINBs were overexpressed in peri-implant tissues and peri-implantitis patients with PICF. SERPINBs was positively correlated to IL-6 and TNF-α in peri-implantitis patients with PICF. The ROC-AUCs of SERPINBs achieved a significantly higher range from 0.895 to 0.939 in peri-implantitis patients with PICF. Therefore, certain SERPINBs expressions were not only perceived through PICF and peri-implant tissues but also showed potential significance in peri-implantitis. CONCLUSION: SERPINBs play an influential role in the pathogenesis of peri-implantitis via binding with other inflammatory cytokines.

Genome-based studies indicate that the Enterococcus faecium Clade B strains belong to Enterococcus lactis species and lack of the hospital infection associated markers.

Enterococcus lactis and the heterotypic synonym Enterococcus xinjiangensis from dairy origin have recently been identified as a novel species based on 16S rRNA gene sequence analysis. Enterococcus faecium type strain NCTC 7171T was used as the reference genome for determining E. lactis and E. faecium to be separate species. However, this taxonomic classification did not consider the diverse lineages of E. faecium, and the double nature of hospital-associated (clade A) and community-associated (clade B) isolates. Here, we investigated the taxonomic relationship among isolates of E. faecium of different origins and E. lactis, using a genome-based approach. Additional to 16S rRNA gene sequence analysis, we estimated the relatedness among strains and species using phylogenomics based on the core pangenome, multilocus sequence typing, the average nucleotide identity and digital DNA-DNA hybridization. Moreover, following the available safety assessment schemes, we evaluated the virulence profile and the ampicillin resistance of E. lactis and E. faecium clade B strains. Our results confirmed the genetic and evolutionary differences between clade A and the intertwined clade B and E. lactis group. We also confirmed the absence in these strains of virulence gene markers IS16, hylEfm and esp and the lack of the PBP5 allelic profile associated with ampicillin resistance. Taken together, our findings support the reassignment of the strains of E. faecium clade B as E. lactis.

Virological Characterization of Critically Ill Patients With COVID-19 in the United Kingdom: Interactions of Viral Load, Antibody Status, and B.1.1.7 Infection.

BACKGROUND: Convalescent plasma containing neutralizing antibody to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is under investigation for coronavirus disease 2019 (COVID-19) treatment. We report diverse virological characteristics of UK intensive care patients enrolled in the Immunoglobulin Domain of the REMAP-CAP randomized controlled trial that potentially influence treatment outcomes. METHODS: SARS-CoV-2 RNA in nasopharyngeal swabs collected pretreatment was quantified by PCR. Antibody status was determined by spike-protein ELISA. B.1.1.7 was differentiated from other SARS-CoV-2 strains using allele-specific probes or restriction site polymorphism (SfcI) targeting D1118H. RESULTS: Of 1274 subjects, 90% were PCR positive with viral loads 118-1.7 × 1011IU/mL. Median viral loads were 40-fold higher in those IgG seronegative (n = 354; 28%) compared to seropositives (n = 939; 72%). Frequencies of B.1.1.7 increased from <1% in November 2020 to 82% of subjects in January 2021. Seronegative individuals with wild-type SARS-CoV-2 had significantly higher viral loads than seropositives (medians 5.8 × 106 and 2.0 × 105 IU/mL, respectively; P = 2 × 10-15). CONCLUSIONS: High viral loads in seropositive B.1.1.7-infected subjects and resistance to seroconversion indicate less effective clearance by innate and adaptive immune responses. SARS-CoV-2 strain, viral loads, and antibody status define subgroups for analysis of treatment efficacy.

Antiretroviral Imprints and Genomic Plasticity of HIV-1 pol in Non-clade B: Implications for Treatment.

Combinational antiretroviral therapy (cART) is the most effective tool to prevent and control HIV-1 infection without an effective vaccine. However, HIV-1 drug resistance mutations (DRMs) and naturally occurring polymorphisms (NOPs) can abrogate cART efficacy. Here, we aimed to characterize the HIV-1 pol mutation landscape in Cameroon, where highly diverse HIV clades circulate, and identify novel treatment-associated mutations that can potentially affect cART efficacy. More than 8,000 functional Cameroonian HIV-1 pol sequences from 1987 to 2020 were studied for DRMs and NOPs. Site-specific amino acid frequencies and quaternary structural features were determined and compared between periods before (≤2003) and after (2004-2020) regional implementation of cART. cART usage in Cameroon induced deep mutation imprints in reverse transcriptase (RT) and to a lower extent in protease (PR) and integrase (IN), according to their relative usage. In the predominant circulating recombinant form (CRF) 02_AG (CRF02_AG), 27 canonical DRMs and 29 NOPs significantly increased or decreased in RT during cART scale-up, whereas in IN, no DRM and only seven NOPs significantly changed. The profound genomic imprints and higher prevalence of DRMs in RT compared to PR and IN mirror the dominant use of reverse transcriptase inhibitors (RTIs) in sub-Saharan Africa and the predominantly integrase strand transfer inhibitor (InSTI)-naïve study population. Our results support the potential of InSTIs for antiretroviral treatment in Cameroon; however, close surveillance of IN mutations will be required to identify emerging resistance patterns, as observed in RT and PR. Population-wide genomic analyses help reveal the presence of selective pressures and viral adaptation processes to guide strategies to bypass resistance and reinstate effective treatment.

An infectious cDNA clone of a growth attenuated Korean isolate of MERS coronavirus KNIH002 in clade B.

The MERS-CoV isolated during the 2015 nosocomial outbreak in Korea showed distinctive differences in mortality and transmission patterns compared to the prototype MERS-CoV EMC strain belonging to clade A. We established a BAC-based reverse genetics system for a Korean isolate of MERS-CoV KNIH002 in the clade B phylogenetically far from the EMC strain, and generated a recombinant MERS-CoV expressing red fluorescent protein. The virus rescued from the infectious clone and KNIH002 strain displayed growth attenuation compared to the EMC strain. Consecutive passages of the rescued virus rapidly generated various ORF5 variants, highlighting its genetic instability and calling for caution in the use of repeatedly passaged virus in pathogenesis studies and for evaluation of control measures against MERS-CoV. The infectious clone for the KNIH002 in contemporary epidemic clade B would be useful for better understanding of a functional link between molecular evolution and pathophysiology of MERS-CoV by comparative studies with EMC strain.

Diversity of mitochondrial genes and predominance of Clade B in different head lice populations in the northwest of Iran.

BACKGROUND: The head louse, Pediculus humanus capitis, is the most important ectoparasite causing many health problems. Several linkages are presented for this parasite, each representing a particular geographical distribution, prevalence rate, vector competence, susceptibility to pediculicides, and infestation rate. Determining the genetic nature of these linkages is necessary to identify the population structure and also to develop and monitor control programmes against head lice. This study was designed to analyse cox1 and cytb genes and determine the mitochondrial clades in head lice populations in the northwest of Iran. METHODS: Adult head lice were collected from infested females of Ardabil, East and West Azerbaijan, and Zanjan Provinces from 2016 to 2018. Partial fragments of the mitochondrial genes cox1 and cytb were amplified by PCR and some of the amplicons were sequenced. All confirmed sequences were analysed, and the frequency of each mitochondrial clade was determined in the studied areas. RESULTS: A total of 6410 females were clinically examined, and 897 adult head lice were collected from 562 infested cases. Genomic DNA was extracted from 417 samples, and fragments of cox1 and cytb genes were amplified in 348 individuals. Analysis of the 116 sequences showed the 632-bp and 495-bp fragments for cox1 and cytb genes, respectively. The nucleotide and haplotype diversities of cytb and cox1 genes were 0.02261 and 0.589 and 0.01443 and 0.424, respectively. Sequence analysis indicated 6 haplotypes clustered in two clades, A and B. The relative prevalence of clade B was 73% for cytb and 82% for cox1 gene. Haplotypes of clade B were found in all the studied areas, while those of clade A were observed only in rural and suburban areas. CONCLUSIONS: To our knowledge, this is the first study investigated deeply the field populations of Pediculus and documented two clades in the Middle East. The considerable prevalence of pediculosis in the studied areas requires authorities' attention to establish effective control and preventive measures. Given the role of cytb in monitoring population groups, application of this marker is suggested for future epigenetic studies to evaluate the factors affecting the abundance of these clades.

Pythium subutonaiense, A New Aquatic Oomycete from Southern China Based on Morphological and Molecular Characters.

A new species, Pythium subutonaiense, isolated from aquatic environments (lake) in China is being described based on morphological characters and molecular evidence. The isolates grew at temperatures between 5 °C and 38 °C, and the optimum temperature was 30 °C, with a radial growth rate of 17.6 mm at 25 °C per day. It is homothallic and characterized by globose to sub-globose shaped and mostly terminal or sometimes catenulate hyphal swellings, filamentous non-inflated sporangia, and smooth oogonia with hypogynous and monoclinous antheridia that contained one plerotic oospore. In phylogenetic analysis, inferred based on the internal transcribed spacer region of the ribosomal RNA gene and mitochondrial cytochrome c oxidase subunit 1 gene, the new species formed a distinct lineage in Pythium clade B. Differences between the new species and phylogenetically related and morphologically similar species are discussed.

Development and analytical validation of a group-specific RT-qPCR assay for the detection of the Simbu serogroup orthobunyaviruses.

The Simbu serogroup within the genus Orthobunyavirus belongs to the family Peribunyaviridae and comprises 32 recognised three-segmented negative-sense single-stranded RNA viruses, with a cosmopolitan distribution. This group of arthropod-borne viruses includes important pathogens of humans and domestic animals e.g. Oropouche orthobunyavirus and Schmallenberg virus. Sensitive and specific diagnostic tools are required for recognition and control of outbreaks. A novel TaqMan® RT-qPCR assay was developed, optimised and analytically validated for the broad detection of the Simbu serogroup orthobunyaviruses. A region in the S segment, which encodes the nucleocapsid protein, was used to design a group primer set and a pair of differently labelled TaqMan® minor groove binder probes to distinguish phylogenetic clade A and B of the serogroup. Efficiencies determined for seven members of the group were 99% for Akabane orthobunyavirus (AKAV), 96% for Simbu orthobunyavirus (SIMV), 96% for Shuni orthobunyavirus (SHUV), 97% for Sathuperi orthobunyavirus (SATV), 84% for Shamonda orthobunyavirus (SHAV), 93% for Ingwavuma virus (INGV, now classified as Manzanilla orthobunyavirus) and 110% for Sabo virus (SABOV, now classified as AKAV). The 95% limit of detection (TCID50/reaction) was 10-3.61 for AKAV, 10-2.38 for SIMV, 10-3.42 for SHUV, 10-3.32 for SATV, 10-1.67 for SHAV, 100.39 for INGV and 10-2.70 for SABOV.

Epithelial SERPINB10, a novel marker of airway eosinophilia in asthma, contributes to allergic airway inflammation.

Serine peptidase inhibitor, clade B, member 10 (SERPINB10) expression is increased in IL-13-stimulated human bronchial epithelial cells and in a murine model of allergic airway inflammation. However, the role of SERPINB10 in asthma remains unknown. We examined the association between epithelial SERPINB10 expression and airway eosinophilia in subjects with asthma and the role of Serpinb10 in allergic airway inflammation in an animal model. Epithelial SERPINB10 mRNA and protein expression were markedly increased in subjects with asthma ( n = 60) compared with healthy controls ( n = 25). Epithelial SERPINB10 mRNA levels were significantly correlated with airway hyperresponsiveness (AHR) and three parameters reflecting airway eosinophilia including the percentage of sputum eosinophils, the number of eosinophils in bronchial submucosa, and fraction of exhaled nitric oxide in subjects with asthma. Moreover, epithelial SERPINB10 expression was strongly correlated with the epithelial gene signature ( CLCA1, POSTN, and SERPINB2) for type 2 status. In normal human bronchial epithelial cells cultured at air-liquid interface, knockdown of SERPINB10 suppressed IL-13-stimulated periostin (encoded by POSTN) and CCL26 (eotaxin-3) expression by inhibiting the activation of p38 MAPK. Epithelial CCL26 mRNA levels were correlated with SERPINB10 expression in subjects with asthma. Airway knockdown of Serpinb10 alleviated AHR, airway eosinophilia and the expression of periostin and Ccl26 in a murine model of allergic airway disease. Taken together, epithelial SERPINB10 is a novel marker for airway eosinophilia in asthma. Epithelial SERPINB10 contributes to allergic airway eosinophilic inflammation, at least in part, by regulating the expression of periostin and CCL26.

New Species of Closely Related Endosymbiotic Dinoflagellates in the Greater Caribbean have Niches Corresponding to Host Coral Phylogeny.

Symbiotic dinoflagellates in the genus Breviolum (formerly Symbiodinium Clade B) dominate coral communities in shallow waters across the Greater Caribbean. While some formally described species exist, mounting genetic, and ecological evidence indicate that numerous more comprise this genus, many of which are closely related. To test this, colonies of common reef-building corals were sampled across a large geographical range. Phylogenetic and population genetic markers then used to examine evolutionary divergence and delineate boundaries of genetic recombination. Three new candidate species were distinguished by fixed differences in nucleotide sequences from nuclear and chloroplast DNA. Population connectivity was evident within each lineage over thousands of kilometers, however, substantial genetic structure persisted between lineages co-occurring within sampling locations, signifying reproductive isolation. While geographically widespread with overlapping distributions, each species is ecologically distinct, exhibiting specific mutualisms with phylogenetically distinct coral hosts. Moreover, significant differences in mean cell sizes provide some morphological evidence substantiating formal species distinctions. In providing evidence that satisfies the biological, phylogenetic, ecological, and morphological species concepts, we classify and formally name Breviolum faviinorum n. sp., primarily associated with Caribbean corals belonging to the Caribbean subfamily Faviinae; B. meandrinium n. sp., associated with corals belonging to the family Meandrinidae; and B. dendrogyrum n. sp., a symbiont harbored exclusively by the threatened coral Dendrogyra cylindrus. These findings support the primary importance of niche diversification (i.e. host habitat) in the speciation of symbiotic dinoflagellates.

An updated assessment of Symbiodinium spp. that associate with common scleractinian corals from Moorea (French Polynesia) reveals high diversity among background symbionts and a novel finding of clade B.

The adaptative bleaching hypothesis (ABH) states that, depending on the symbiotic flexibility of coral hosts (i.e., the ability of corals to "switch" or "shuffle" their algal symbionts), coral bleaching can lead to a change in the composition of their associated Symbiodinium community and, thus, contribute to the coral's overall survival. In order to determine the flexibility of corals, molecular tools are required to provide accurate species delineations and to detect low levels of coral-associated Symbiodinium. Here, we used highly sensitive quantitative (real-time) PCR (qPCR) technology to analyse five common coral species from Moorea (French Polynesia), previously screened using only traditional molecular methods, to assess the presence of low-abundance (background) Symbiodinium spp. Similar to other studies, each coral species exhibited a strong specificity to a particular clade, irrespective of the environment. In addition, however, each of the five species harboured at least one additional Symbiodinium clade, among clades A-D, at background levels. Unexpectedly, and for the first time in French Polynesia, clade B was detected as a coral symbiont. These results increase the number of known coral-Symbiodinium associations from corals found in French Polynesia, and likely indicate an underestimation of the ability of the corals in this region to associate with and/or "shuffle" different Symbiodinium clades. Altogether our data suggest that corals from French Polynesia may favor a trade-off between optimizing symbioses with a specific Symbiodinium clade(s), maintaining associations with particular background clades that may play a role in the ability of corals to respond to environmental change.

Differences in Glycoprotein Complex Receptor Binding Site Accessibility Prompt Poor Cross-Reactivity of Neutralizing Antibodies between Closely Related Arenaviruses.

The glycoprotein complex (GPC) of arenaviruses, composed of stable signal peptide, GP1, and GP2, is the only antigen correlated with antibody-mediated neutralization. However, despite strong cross-reactivity of convalescent antisera between related arenavirus species, weak or no cross-neutralization occurs. Two closely related clade B viruses, Machupo virus (MACV) and Junín virus (JUNV), have nearly identical overall GPC architecture and share a host receptor, transferrin receptor 1 (TfR1). Given structural and functional similarities of the GP1 receptor binding site (RBS) of these viruses and the recent demonstration that the RBS is an important target for neutralizing antibodies, it is not clear how these viruses avoid cross-neutralization. To address this, MACV/JUNV chimeric GPCs were assessed for interaction with a group of α-JUNV GPC monoclonal antibodies (MAbs) and mouse antisera against JUNV or MACV GPC. All six MAbs targeted GP1, with those that neutralized JUNV GPC-pseudovirions competing with each other for RBS binding. However, these MAbs were unable to bind to a chimeric GPC composed of JUNV GP1 containing a small disulfide bonded loop (loop 10) unique to MACV GPC, suggesting that this loop may block MAbs interaction with the GP1 RBS. Consistent with this loop causing interference, mouse anti-JUNV GPC antisera that solely neutralized pseudovirions bearing autologous GP1 provided enhanced neutralization of MACV GPC when this loop was removed. Our studies provide evidence that loop 10, which is unique to MACV GP1, is an important impediment to binding of neutralizing antibodies and contributes to the poor cross-neutralization of α-JUNV antisera against MACV.IMPORTANCE Multiple New World arenaviruses can cause severe disease in humans, and some geographic overlap exists among these viruses. A vaccine that protects against a broad range of New World arenaviruses is desirable for purposes of simplicity, cost, and broad protection against multiple National Institute of Allergy and Infectious Disease-assigned category A priority pathogens. In this study, we sought to better understand how closely related arenaviruses elude cross-species neutralization by investigating the structural bases of antibody binding and avoidance. In our studies, we found that neutralizing antibodies against two New World arenaviruses, Machupo virus (MACV) and Junín virus (JUNV), bound to the envelope glycoprotein 1 (GP1) with JUNV monoclonal antibodies targeting the receptor binding site (RBS). We further show that altered structures surrounding the RBS pocket in MACV GP1 impede access of JUNV-elicited antibodies.

Association between different anti-Tat antibody isotypes and HIV disease progression: data from an African cohort.

BACKGROUND: The presence of IgG and IgM against Tat, an HIV protein important for viral replication and immune dysfunction, is associated with slow disease progression in clade B HIV-infected individuals. However, although Tat activities strictly depend on the viral clade, our knowledge about the importance of anti-Tat antibodies in non-clade B HIV infection is poor. The objective of this study was to investigate the association of different anti-Tat antibody isotypes with disease progression in non-clade B HIV-infected subjects and to study the relationship between anti-Tat humoral responses and immunological abnormalities. METHODS: Anti-clade B and -clade C Tat IgG, IgM and IgA titers were assessed in serum samples from 96 cART-naïve subjects with chronic HIV infection from Mbeya, Tanzania, and associated with CD4(+) T cell count, plasma viremia and CD4(+) and CD8(+) T cell phenotypes. RESULTS: Anti-Tat IgM were preferentially detected in chronic HIV-infected subjects with low T cell activation (p-value = 0.03) and correlated with higher CD4(+) T cell counts and lower viral loads irrespective of the duration of infection (p-value = 0.019 and p-value = 0.037 respectively). Conversely, anti-Tat IgA were preferentially detected in individuals with low CD4(+) T cell counts and high viral load (p-value = 0.02 and p-value < 0.001 respectively). The simultaneous presence of anti-Tat IgG and IgM protected from fast CD4(+) T cell decline (p-value < 0.01) and accumulation of CD38(+)HLADR(+)CD8(+) T cells (p- value = 0.029). CONCLUSIONS: Anti-Tat IgG alone are not protective in non-clade B infected subjects, unless concomitant with IgM, suggesting a protective role of persistent anti-Tat IgM irrespective of the infecting clade.

Presentation of available CTL epitopes that induction of cell-mediated immune response against HIV-1 Koran clade B strain using computational technology.

OBJECTIVE: Theoretical predicting cytotoxic T lymphocyte (CTL) epitopes are an important tool in vaccine design and CTL therapy for enhancing our understanding of the cellular immune system. We would like to identify available CTL epitopes against HIV-1 Korean clade B. CTL activity was assessed in freshly isolated peripheral blood mononuclear cells from Korean HIV patients in order to assess whether these CTL epitopes induce a cell-mediated immune response (CMI). METHODS: NetCTLpan1.1 software, which is the most popular prediction computer software package, and full atom-based simulation (FABS), which is a 3D modelling system for binding activity between epitopes and human leucocyte antigen (HLA) molecules, were used to predict the peptide-spanning Env region binding to HLA-A*24:02, HLA-A*02:01 and HLA-B*15:01, which are frequently found in the Korean population. Granzyme B and interferon-γ ELISPOT assays were used to determine whether identified CTL epitopes induce CMI. RESULTS: Three HIV-1 Korean clade B-specific Env CTL epitopes were identified: Gp41-RYL and Gp41-RQG are localized within gp41, and Gp120-LLQ within gp120. In in vitro assays using granzyme B ELISPOT, Gp120-LLQ and Gp41-RQG induced epitope-specific CTL responses in HLA-restricted cells. In ex vivo assay using IFN-γ ELISPOT, cell-mediated immune responses to Gp41-RYL were present in 50% of HLA-matched patients, and responses to Gp120-LLQ and Gp41-RQG were found in 33% of HLA-matched patients. CONCLUSION: In this study, we found that a prediction pipeline for CTL epitopes might be based on the most popular computer prediction software and FABS methods. Our results suggest that these CTL epitopes may provide useful tools and information for the development of a therapeutic vaccine against HIV-1 Korean clade B.

Survivorship and feeding preferences among size classes of outplanted sea urchins, Tripneustes gratilla, and possible use as biocontrol for invasive alien algae.

We investigate the survivorship, growth and diet preferences of hatchery-raised juvenile urchins, Tripneustes gratilla, to evaluate the efficacy of their use as biocontrol agents in the efforts to reduce alien invasive algae. In flow-through tanks, we measured urchin growth rates, feeding rates and feeding preferences among diets of the most common invasive algae found in Kane'ohe Bay, Hawai'i: Acanthophora spicifera, Gracilaria salicornia, Eucheuma denticulatum and Kappaphycus clade B. Post-transport survivorship of outplanted urchins was measured in paired open and closed cages in three different reef environments (lagoon, reef flat and reef slope) for a month. Survivorship in closed cages was highest on the reef flat (∼75%), and intermediate in the lagoon and reef slope (∼50%). In contrast, open cages showed similar survivorship on the reef flat and in the lagoon, but only 20% of juvenile urchins survived in open cages placed on the reef slope. Urchins grew significantly faster on diets of G. salicornia (1.58 mm/week ± 0.14 SE) and Kappaphycus clade B (1.69 ± 0.14 mm/wk) than on E. denticulatum (0.97 ± 0.14 mm/wk), with intermediate growth when fed on A. spicifera (1.23 ± 0.11 mm/wk). Interestingly, urchins display size-specific feeding preferences. In non-choice feeding trials, small urchins (17.5-22.5 mm test diameter) consumed G. salicornia fastest (6.08 g/day ± 0.19 SE), with A. spicifera (4.25 ± 0.02 g/day) and Kappaphycus clade B (3.83 ± 0.02 g/day) intermediate, and E. denticulatum was clearly the least consumed (2.32 ± 0.37 g/day). Medium-sized (29.8-43.8 mm) urchins likewise preferentially consumed G. salicornia (12.60 ± 0.08 g/day), with less clear differences among the other species in which E. denticulatum was still consumed least (9.35 ± 0.90 g/day). In contrast, large urchins (45.0-65.0 mm) showed no significant preferences among the different algae species at all (12.43-15.24 g/day). Overall consumption rates in non-choice trials were roughly equal to those in the choice trials, but differences among feeding rates on each species were not predictive of feeding preferences when urchins were presented all four species simultaneously. In the choice feeding trials, both small and medium urchins clearly preferred A. spicifera over all other algae (roughly twice as much consumed as any other species). Again, however, differences were less pronounced among adult urchins, with adults showing a significant preference for A. spicifera and Kappaphycus clade B compared to the other two algal species. These findings indicate that outplanted urchins are surviving on the reef flats and eating a variety of alien invasive algae as intended. Although juvenile urchins show stronger feeding preferences, these animals grow quickly, and adult urchins are more generalist herbivores that consume all four alien invasive algae.