Molecular identification of human enteroviruses in children with neurological infections from the central region of Argentina

In the central area of Argentina, epidemiological and molecular characteristics of human enterovirus infections are still unknown. RT-nested PCR of the highly conserved 5′NCR was used to detect enteroviruses in 168 samples of cerebrospinal fluid from hospitalized patients with suspected infection of the central nervous system (2007–2008), and 13 (7.7%) were positive. Molecular typing was performed by sequencing of the 3′-half VP1 region. Echovirus 30 was the predominant type detected, followed by coxsackie viruses A9 and B4. All echovirus 30 strains of 2007 clustered in lineage H, whereas the echovirus 30 isolate obtained in 2008 was more distantly related, possibly representing a new lineage.     

The Enterprise,a massive transposon carrying Spok meiotic drive genes

The genomes of eukaryotes are full of parasitic sequences known as transposable elements (TEs). Here, we report the discovery of a putative giant tyrosine-recombinase-mobilized DNA transposon, Enterprise, from the model fungus Podospora anserina. Previously, we described a large genomic feature called the Spok block which is notable due to the presence of meiotic drive genes of the Spok gene family. The Spok block ranges from 110 kb to 247 kb and can be present in at least four different genomic locations within P. anserina, despite what is an otherwise highly conserved genome structure. We propose that the reason for its varying positions is that the Spok block is not only capable of meiotic drive but is also capable of transposition. More precisely, the Spok block represents a unique case where the Enterprise has captured the Spoks, thereby parasitizing a resident genomic parasite to become a genomic hyperparasite. Furthermore, we demonstrate that Enterprise (without the Spoks) is found in other fungal lineages, where it can be as large as 70 kb. Lastly, we provide experimental evidence that the Spok block is deleterious, with detrimental effects on spore production in strains which carry it. This union of meiotic drivers and a transposon has created a selfish element of impressive size in Podospora, challenging our perception of how TEs influence genome evolution and broadening the horizons in terms of what the upper limit of transposition may be.

Transposable elements (TEs) are major agents of change in eukaryotic genomes. Their ability to selfishly parasitize their host replication machinery has large impacts on both genome size and on gene regulation (Chénais et al. 2012). In extreme cases, TEs can contribute up to 85% of genomic content (Schnable et al. 2009), and expansion and reduction of TEs can result in rapid changes in both genome size and architecture (Haas et al. 2009; Möller and Stukenbrock 2017; Talla et al. 2017). Generally, TEs have small sizes (∼50–12,000 bp) and accomplish these large-scale changes through their sheer number. For example, there are ∼1.1 million Alu elements in the human genome, which have had a large impact on genome evolution (Jurka 2004; Bennett et al. 2008). The largest known cases among Class I retrotransposons are long terminal repeat (LTR) elements that can be as large as 30 kb, but among Class II DNA transposons, Mavericks/Polintons are known to grow as large as 40 kb through the capture of additional open reading frames (ORFs) (Arkhipova and Yushenova 2019). Recently, a behemoth TE named Teratorn was described in teleost fish; it can be up to 182 kb in length, dwarfing all other known TEs. Teratorn has achieved this impressive size by fusing a piggyBac DNA transposon with a herpesvirus, thereby blurring the line between TEs and viruses (Inoue et al. 2017, 2018). Truly massive transposons may be lurking in the depths of many eukaryotic genomes, but the limitations of short-read genome sequencing technologies and the lack of population-level high-quality assemblies may make them difficult to identify.The Spok block is a large genomic feature that was first identified thanks to the presence of the spore killing (Spok) genes in species from the genus Podospora (Grognet et al. 2014; Vogan et al. 2019). The Spoks are selfish genetic elements that bias their transmission to the next generation in a process known as meiotic drive. Here, drive occurs by inducing the death of spores that do not inherit them, through a single protein that operates as both a toxin and an antidote (Grognet et al. 2014; Vogan et al. 2019). The first Spok gene described, Spok1, was discovered in Podospora comata (Grognet et al. 2014). In P. anserina, the homologous gene Spok2 is found at high population frequencies, whereas two other genes of the Spok family, Spok3 and Spok4, are at low to intermediate frequencies (Vogan et al. 2019). Unlike Spok1 and Spok2, however, Spok3 and Spok4 are always associated with a large genomic region (the Spok block). The Spok block can be located at different chromosomal locations in different individuals but is never found more than once in natural strains. The number of Spok genes and the location of the Spok block (which carries Spok3, Spok4, or both) define the overall meiotic driver behavior of a given genome, which can be classified into the so-called Podospora spore killers or Psks (van der Gaag et al. 2000; Vogan et al. 2019). The Spok block stands out not only because of its size, typically around 150 kb, but also because there is otherwise high genome collinearity among strains of P. anserina and with the related species P. comata and P. pauciseta (Vogan et al. 2019).The fact that the Spok block is found at unique genomic positions between otherwise highly similar strains is of prime interest as each novel Spok block position creates a unique meiotic drive type (Psk) due to the intricacies of meiosis in Podospora (Vogan et al. 2019). We therefore set out to determine the mechanism through which the Spok block relocates throughout the genome. Additionally, we annotated the gene content of the various Spok blocks to describe their composition and understand what represents the minimal component of the Spok block. Lastly, we conducted fitness assays to investigate whether the presence of the Spok block imparts any detrimental effects upon the host.     

Acute gastroenteritis outbreaks in closed and semi-closed facilities during 2017 in Catalonia,Spain

Acute gastroenteritis (AGE) is a leading cause of disease worldwide. The aim of this prospective observational study is to describe the epidemiology of AGE in closed and semi-closed institutions in Catalonia. In 2017, 151 outbreaks were reported; 30.5% occurred in closed and semi-closed institutions; 71.7% caused by norovirus (NoV) (1532) cases. Person-to-person transmission accounted for 75.8% of NoV outbreaks vs 46.1% in non-NoV outbreaks (p?<?0.001). Attack rate for NoV outbreaks was 33.1% vs 14.3% for non-NoV outbreaks (RR?=?2.3; 95%CI: 2.0–2.7). The high number of affected underscores prompt and intense preventive measures to avoid the extension and perpetuation of outbreaks in these settings.

     

Cloning and expression of a truncated form of envelope glycoprotein D of Bovine herpesvirus type 5 in methylotrophic yeast Pichia pastoris

Meningoencephalitis caused by Bovine herpesvirus type 5 (BoHV-5) is responsible for heavy economic losses in the cattle industry. As in other Alphaherpesviruses, the envelope glycoprotein IV (gD), which mediates penetration into host cells, is one of the major candidate antigens for a recombinant vaccine, since it induces a strong and persistent immune response. The DNA coding for a truncated form of BoHV-5 gD (tgD) has been cloned into the Pichia pastoris expression vector pPICZαB to allow protein secretion into the medium. After induction with methanol, a 55 kDa protein was obtained. Enzyme deglycosylation with Endo H showed a smaller size band in SDS-PGAE, with 50 kDa, suggesting that tgD has N-linked oligosaccharides and that it is not hyperglycosylated. The 55 kDa protein was recognized by several polyclonal antibodies, including polyclonal antibody anti-tgD and polyclonal antibodies of different animal species immunized with BoHV-5 and BoHV-1. This is the first report of BoHV-5 gD expression in yeast. It was shown that the recombinant truncated form of BoHV-5 gD has antigenic and immunogenic properties similar to the native BoHV-5 gD. Expression of tgD as a secreted protein allows simple and inexpensive purification methods that can be used for further studies to evaluate its immunogenicity in cattle.     

Banding techniques on tardigrade chromosomes: the karyotype of Macrobiotus richtersi (Eutardigrada, Macrobiotidae)

This work represents the first attempt to define tardigrade chromosomes using banding techniques. Macrobiotus richtersi, a eutardigrade morphospecies with amphimictic diploid and thelytokous triploid cytotypes, was used as a model. Prime consideration was given to oocyte chromosomes because they are larger than those of spermatocytes and of mitotic chromosomes. With Giemsa staining, the chromatids of the 6 bivalents of the diploid cytotypes and those of the 17–18 univalents of the triploid cytotypes were very similar to each other and appeared rod- or flame-shaped. In the amphimictic strain, a chiasma was generally present in each bivalent at diplotene, whereas there were no chiasmata in the oocyte prophase chromosomes of the triploid strain. Both in diploid and triploid cytotypes, C-banding and fluorescence showed a heterochromatic centromeric band on the telomere of each chromosome oriented towards the spindle pole, indicating that all of them were acrocentric. Silver staining showed the presence of a NOR in only a pair of chromosomes, close to the centromeric C-banded site. NOR was particularly evident in the oocyte prophases. Other silver positive regions, corresponding to the kinetochore, were located on all other chromosomes on the telomeres towards the spindle pole. This revised version was published online in July 2006 with corrections to the Cover Date.     

Different patterns of 11q allelic losses in digestive endocrine tumors

Most foregut digestive endocrine neoplasms may be associated with the multiple endocrine type 1 (MEN-1) syndrome. In contrast, midgut/hindgut carcinoids never show such association. To investigate the pathogenetic involvement of the MEN-1 gene and of putative additional oncosuppressor gene(s) distal to it, a comparative analysis of loss of heterozygosity (LOH) at chromosome 11q13 to 11qter was performed in 27 foregut (pancreatic endocrine tumors [PETs]), 23 midgut (ileal and appendiceal), and 3 hindgut (rectal) endocrine tumors. LOH at the MEN-1 gene locus at 11q13 was observed in 52% of the 23 sporadic and in all 4 MEN-1-associated PETs and was found to consistently and continuously span to the most distal marker investigated at 11qter. In contrast, only occasional, discontinuous, and mostly interstitial LOH for 11q markers was observed in ileal (midgut) carcinoids, whereas no LOH was found in all appendiceal (midgut) and rectal (hindgut) carcinoids. The consistent extension of LOH from the MEN-1 region to 11qter in sporadic PETs suggests a mechanism of gene inactivation via chromosomal breakage and complete loss of chromosome 11q; furthermore, these results expand beyond the 11q13 region the search for additional oncosuppressor gene(s) potentially involved in the genesis of these neoplasms. The low frequency, limited extension, and discontinuous distribution of 11q deletions in midgut/hindgut carcinoids suggest that MEN-1 gene is not involved in the pathogenesis of these tumors.     

Simple Synthesis of Aldehyde and Carboxylic Acid Terminated Methoxypoly(ethylene glycol)

mPEG‐aldehydes and activated mPEG‐carboxylic acids on which the polymer and the reactive functional group are separated by aliphatic chains of different lengths, are well known reagents of widespread application in PEGylation technology. The syntheses of these functional polymers are not easy and usually the preparation of each of these compounds requires a very specific set of reagents and experimental conditions. In fact, a general method for their preparation is, still nowadays, unavailable. In this work a simple synthetic method is developed for preparation of mPEG‐aldehydes and carboxylic acids with excellent yields. The key step of the method involves a substitution reaction of mPEG‐mesylate with mono‐alkoxides of symmetrical diols to afford hydroxyl‐terminated polymers, which are converted to the aldehydes and acids by mild oxidation reactions.

     

Increased lipid and protein oxidation and lowered anti-oxidant defenses in systemic lupus erythematosus are associated with severity of illness,autoimmunity, increased adhesion molecules,and Th1 and Th17 immune shift

This study investigated nitro-oxidative stress in patients with systemic lupus erythematosus (SLE) in association with disease activity, immune-inflammatory biomarkers, and adhesion molecules. Two-hundred-four patients with SLE and 256 healthy volunteers were enrolled in this case-control study, which measured nitro-oxidative stress biomarkers, including lipid peroxides (LOOH), advanced oxidation protein products (AOPPs), nitric oxide metabolites (NO_x), sulfhydryl (?SH) groups, products of deoxyribonucleic acid (DNA)/ribonucleic acid (RNA) oxidative degradation, and total radical-trapping anti-oxidant parameter (TRAP). Also measured were anti-nuclear antibodies (ANAs), antibodies against double-stranded DNA (dsDNA), plasma levels of diverse cytokines, C-reactive protein, and adhesion molecules. LOOH (p < 0.001) and AOPP (p < 0.001) were significantly higher, while TRAP was significantly lower (p < 0.001) in SLE patients than in controls. AOPP and LOOH were significantly and positively associated with SLE disease activity index (SLEDAI) scores, anti-nuclear antibodies, and antibodies against double-stranded DNA (anti-dsDNA) levels, while TRAP was significantly and inversely correlated with SLEDAI, ANA, and dsDNA antibody levels. There were significant positive associations between AOPP and LOOH and immune-inflammatory markers, indicating T helper (Th)-17 and Th1 bias and Th1 + Th17/Th2 ratio (p = 0.002 and p = 0.001, respectively). AOPP and LOOH (positively) and TRAP (inversely) were associated with adhesion molecule expression. A model predicting SLE was computed showing that, using LOOH, AOPP, NO_x, adhesion molecules, and body mass index, 94.2% of the patients were correctly classified with a specificity of 91.5%. Increased nitro-oxidative stress takes part in the (auto)immune pathophysiology of SLE and modulates severity of illness and adhesion molecule expression.