Roberto Bertorelli

HIV-1 Nef promotes infection by excluding SERINC5 from virion incorporation

HIV-1 Nef, a protein important for the development of AIDS, has well-characterized effects on host membrane trafficking and receptor downregulation. By an unidentified mechanism, Nef increases the intrinsic infectivity of HIV-1 virions in a host-cell-dependent manner. Here we identify the host transmembrane protein SERINC5, and to a lesser extent SERINC3, as a potent inhibitor of HIV-1 particle infectivity that is counteracted by Nef. SERINC5 localizes to the plasma membrane, where it is efficiently incorporated into budding HIV-1 virions and impairs subsequent virion penetration of susceptible target cells. Nef redirects SERINC5 to a Rab7-positive endosomal compartment and thereby excludes it from HIV-1 particles. The ability to counteract SERINC5 was conserved in Nef encoded by diverse primate immunodeficiency viruses, as well as in the structurally unrelated glycosylated Gag from murine leukaemia virus. These examples of functional conservation and convergent evolution emphasize the fundamental importance of SERINC5 as a potent anti-retroviral factor.

Unexplored diversity and strain-level structure of the skin microbiome associated with psoriasis

Psoriasis is an immune-mediated inflammatory skin disease that has been associated with cutaneous microbial dysbiosis by culture-dependent investigations and rRNA community profiling. We applied, for the first time, high-resolution shotgun metagenomics to characterise the microbiome of psoriatic and unaffected skin from 28 individuals. We demonstrate psoriatic ear sites have a decreased diversity and psoriasis is associated with an increase in Staphylococcus, but overall the microbiomes of psoriatic and unaffected sites display few discriminative features at the species level. Finer strain-level analysis reveals strain heterogeneity colonisation and functional variability providing the intriguing hypothesis of psoriatic niche-specific strain adaptation or selection. Furthermore, we accessed the poorly characterised, but abundant, clades with limited sequence information in public databases, including uncharacterised Malassezia spp. These results highlight the skins hidden diversity and suggests strain-level variations could be key determinants of the psoriatic microbiome. This illustrates the need for high-resolution analyses, particularly when identifying therapeutic targets. This work provides a baseline for microbiome studies in relation to the pathogenesis of psoriasis.Psoriasis: investigating microbial diversityAnalysing microbial populations on the skin provides an insight into the diversity of microbes associated with psoriasis. Nicola Segata and colleagues at the University of Trento, Italy, used genetic analysis to compare the microbial populations on regions of skin affected and unaffected by psoriasis. Staphylococcus bacteria were more prevalent in psoriasis, but there was little clearly defined difference in microbial species on psoriasis-affected and unaffected skin. There was, however, decreased microbial diversity on psoriatic ear sites. Deeper strain-level computational analysis suggested that psoriasis could offer niche locations for colonisation by specific strains of staphylococci and propionibacteria. The results highlight the diversity of microbial populations on the skin, and the need for larger cohorts to build on the baseline data now established. Further studies might help identify targets for treating skin bacteria associated with psoriasis.

Genomic characterization of Nontuberculous Mycobacteria

Mycobacterium tuberculosis and Mycobacterium leprae have remained, for many years, the primary species of the genus Mycobacterium of clinical and microbiological interest. The other members of the genus, referred to as nontuberculous mycobacteria (NTM), have long been underinvestigated. In the last decades, however, the number of reports linking various NTM species with human diseases has steadily increased and treatment difficulties have emerged. Despite the availability of whole genome sequencing technologies, limited effort has been devoted to the genetic characterization of NTM species. As a consequence, the taxonomic and phylogenetic structure of the genus remains unsettled and genomic information is lacking to support the identification of these organisms in a clinical setting. In this work, we widen the knowledge of NTMs by reconstructing and analyzing the genomes of 41 previously uncharacterized NTM species. We provide the first comprehensive characterization of the genomic diversity of NTMs and open new venues for the clinical identification of opportunistic pathogens from this genus.

Characterization of 17 strains belonging to the Mycobacterium simiae complex and description of Mycobacterium paraense sp. nov.

Fourteen mycobacterial strains isolated from pulmonary samples of independent patients in the state of Pará (Brazil), and three strains isolated in Italy, were characterized using a polyphasic approach. Thorough genetic investigation, including whole-genome sequencing, demonstrated that the strains belong to the M. simiae complex, being most closely related to Mycobacterium interjectum. For 14 of the strains, evidence emerged supporting their inclusion in a previously unreported species of the genus Mycobacterium, for which the name Mycobacterium paraense sp. nov. is proposed (type strain, IEC26(T) = DSM 46749(T) = CCUG 66121(T)). The novel species is characterized by slow growth, unpigmented or pale yellow scotochromogenic colonies, and a HPLC mycolic acid profile different from other known mycobacteria. In different genetic regions, high sequence microheterogeneity was detected.

Enhanced microbial diversity in the saliva microbiome induced by short-term probiotic intake revealed by 16S rRNA sequencing on the IonTorrent PGM platform.

Microbial communities populating several human body habitats are important determinants of human health. Cultivation-free community-wide approaches like bacterial 16S rRNA sequencing recently revolutionized the study of such human-associated microbial diversity, and the continuously decreasing cost/throughput ratio of current sequencing platforms is further enhancing the availability and effectiveness of microbiome research. The IonTorrent PGM platform is among the latest available commercial high-throughput sequencing tools, but it is just starting to be used for 16S rRNA surveys with only episodic assessments of its performance. We present here the first IonTorrent profiling of the human saliva microbiome collected from 12 healthy individuals. In this cohort, a subset of the volunteers was asked to assume a probiotic product, in order to investigate its impact on the composition and the structure of the saliva microbiome. Analysis of the generated dataset suggests the suitability of the IonTorrent platform for 16S rRNA surveys, even though some platform-specific choices are required to optimize the consistency of the obtained bacterial profiles. Interestingly, we found a marked and statistically significant increase of the overall bacterial diversity in the saliva of individuals who received the probiotic product compared to the control group, suggesting a short-term effect of probiotic product administration on oral microbiome composition.

The new phylogeny of the genus Mycobacterium: The old and the news

BACKGROUND Phylogenetic studies of bacteria have been based so far either on a single gene (usually the 16S rRNA) or on concatenated housekeeping genes. For what concerns the genus Mycobacterium these approaches support the separation of rapidly and slowly growing species and the clustering of most species in well-defined phylogenetic groups. The advent of high-throughput shotgun sequencing leads us to revise conventional taxonomy of mycobacteria on the light of genomic data. For this purpose we investigated 88 newly sequenced species in addition to 60 retrieved from GenBank and used the Average Nucleotide Identity pairwise scores to reconstruct phylogenetic relationships within this genus. RESULTS Our analysis confirmed the separation of slow and rapid growers and the intermediate position occupied by the M. terrae complex. Among the rapid growers, the species of the M. chelonae-abscessus complex belonged to the most ancestral cluster. Other major clades of rapid growers included the species related to M. fortuitum and M. smegmatis and a large grouping containing mostly environmental species rarely isolated from humans. The members of the M. terrae complex appeared as the most ancestral slow growers. Among slow growers two deep branches led to the clusters of species related to M. celatum and M. xenopi and to a large group harboring most of the species more frequently responsible of disease in humans, including the major pathogenic mycobacteria (M. tuberculosis, M. leprae, M. ulcerans). The species previously grouped in the M. simiae complex were allocated in a number of sub-clades; of them, only the one including the species M. simiae identified the real members of this complex. The other clades included also species previously not considered related to M. simiae. The ANI analysis, in most cases supported by Genome to Genome Distance and by Genomic Signature-Delta Difference, showed that a number of species with standing in literature were indeed synonymous. CONCLUSIONS Genomic data revealed to be much more informative in comparison with phenotype. We believe that the genomic revolution enabled by high-throughput shotgun sequencing should now be considered in order to revise the conservative approaches still informing taxonomic sciences.

Mother-to-Infant Microbial Transmission from Different Body Sites Shapes the Developing Infant Gut Microbiome

Summary The acquisition and development of the infant microbiome are key to establishing a healthy host-microbiome symbiosis. The maternal microbial reservoir is thought to play a crucial role in this process. However, the source and transmission routes of the infant pioneering microbes are poorly understood. To address this, we longitudinally sampled the microbiome of 25 mother-infant pairs across multiple body sites from birth up to 4 months postpartum. Strain-level metagenomic profiling showed a rapid influx of microbes at birth followed by strong selection during the first few days of life. Maternal skin and vaginal strains colonize only transiently, and the infant continues to acquire microbes from distinct maternal sources after birth. Maternal gut strains proved more persistent in the infant gut and ecologically better adapted than those acquired from other sources. Together, these data describe the mother-to-infant microbiome transmission routes that are integral in the development of the infant microbiome.

The CDKN2A/p16INK4a 5′UTR sequence and translational regulation: impact of novel variants predisposing to melanoma

Many variants of uncertain functional significance in cancer susceptibility genes lie in regulatory regions, and clarifying their association with disease risk poses significant challenges. We studied 17 germline variants (nine of which were novel) in the CDKN2A 5′UTR with independent approaches, which included mono and bicistronic reporter assays, Western blot of endogenous protein, and allelic representation after polysomal profiling to investigate their impact on CDKN2A mRNA translation regulation. Two of the novel variants (c.‐27del23, c.‐93‐91delAGG) were classified as causal mutations (score ≥3), along with the c.‐21C>T, c.‐34G>T, and c.‐56G>T, which had already been studied by a subset of assays. The novel c.‐42T>A as well as the previously described c.‐67G>C were classified as potential mutations (score 1 or 2). The remaining variants (c.‐14C>T, c.‐20A>G, c.‐25C>T+c.‐180G>A, c.‐30G>A, c.‐40C>T, c.‐45G>A, c.‐59C>G, c.‐87T>A, c.‐252A>T) were classified as neutral (score 0). In conclusion, we found evidence that nearly half of the variants found in this region had a negative impact on CDKN2A mRNA translation, supporting the hypothesis that 5′UTR can act as a cellular Internal Ribosome Entry Site (IRES) to modulate p16INK4a translation.

Exome sequencing in a patient with Catel–Manzke-like syndrome excludes the involvement of the known genes and reveals a possible candidate

In the present study we describe the exome sequencing and analysis of a patient with Catel-Manzke-like phenotype showing bilateral hyperphalangism of the second finger and thumb clinodactyly due to a unilateral delta phalanx, associated with growth, cardiac and vertebral defects. The exome sequencing analysis excluded pathogenetic mutations in the genes known to cause syndromes with hyperphalangism and did not identify any alteration in the X-chromosome or de novo mutations in likely candidate genes. Under the assumption of an autosomal recessive mode of inheritance and based on the frequency of the single nucleotide variants found in homozygous or double heterozygous states and the results of computer prediction programs, only one gene, DNAH10, emerged as a candidate in the pathogenesis of the disease in our patient. However, the differences among the known biological functions of DNAH10 and the genes involved in the other syndromes with hyperphalangism, suggest caution in the interpretation of the results.

The short-term impact of probiotic consumption on the oral cavity microbiome

The dysbiosis of the oral microbiome is associated with both localized and systemic diseases. Modulating the resident microbial communities by the dietary consumption of probiotics has become an appealing means to promote host health by either restoring host-microbe balance or preventing dysbiosis. Most probiotics strategies target the intestinal microbiome, but little is known about their impact on the oral microbiome. We analyzed here the saliva microbiome from 21 volunteers, longitudinally collected before, during, and after consumption of a commercial probiotic and a standard yoghurt using 16S amplicon sequencing. The alpha diversity of the saliva microbiome had a statistically significant increase (P-value = 0.0011) in one of the groups that consumed the probiotic. The overall structure of the microbiome was however not significantly impacted by the probiotic, although oligotyping analysis revealed that both Streptococci and Lactobacilli present in the probiotic product persisted in the saliva microbiome. In contrast, non-probiotic yoghurt consumption had a lesser impact on the overall diversity and Lactobacillus and Streptococcus persistence. Our results suggest that consumption of commercial probiotics in healthy subjects increase the overall diversity of the oral cavity microbiome in the short term, but such dietary interventions are not able to substantially modify the structure of the microbiome.