Phylogenomics and analysis of shared genes suggest a single transition to mutualism in Wolbachia of nematodes

Wolbachia, endosymbiotic bacteria of the order Rickettsiales, are widespread in arthropods but also present in nematodes. In arthropods, A and B supergroup Wolbachia are generally associated with distortion of host reproduction. In filarial nematodes, including some human parasites, multiple lines of experimental evidence indicate that C and D supergroup Wolbachia are essential for the survival of the host, and here the symbiotic relationship is considered mutualistic. The origin of this mutualistic endosymbiosis is of interest for both basic and applied reasons: How does a parasite become a mutualist? Could intervention in the mutualism aid in treatment of human disease? Correct rooting and high-quality resolution of Wolbachia relationships are required to resolve this question. However, because of the large genetic distance between Wolbachia and the nearest outgroups, and the limited number of genomes so far available for large-scale analyses, current phylogenies do not provide robust answers. We therefore sequenced the genome of the D supergroup Wolbachia endosymbiont of Litomosoides sigmodontis, revisited the selection of loci for phylogenomic analyses, and performed a phylogenomic analysis including available complete genomes (from isolates in supergroups A, B, C, and D). Using 90 orthologous genes with reliable phylogenetic signals, we obtained a robust phylogenetic reconstruction, including a highly supported root to the Wolbachia phylogeny between a (A + B) clade and a (C + D) clade. Although we currently lack data from several Wolbachia supergroups, notably F, our analysis supports a model wherein the putatively mutualist endosymbiotic relationship between Wolbachia and nematodes originated from a single transition event

'Candidatus Sarmatiella mevalonica' endosymbiont of the ciliate Paramecium provides insights on evolutionary plasticity among Rickettsiales

Members of the bacterial order Rickettsiales are obligatorily associated with a wide range of eukaryotic hosts. Their evolutionary trajectories, in particular concerning the origin of shared or differential traits among distant sub-lineages, are still poorly understood. Here, we characterized a novel Rickettsiales bacterium associated with the ciliate Paramecium tredecaurelia and phylogenetically related to the Rickettsia genus. Its genome encodes significant lineage-specific features, chiefly the mevalonate pathway gene repertoire, involved in isoprenoid precursor biosynthesis. Not only this pathway has never been described in Rickettsiales, it also is very rare among bacteria, though typical in eukaryotes, thus likely representing a horizontally acquired trait. The presence of these genes could enable an efficient exploitation of host-derived intermediates for isoprenoid synthesis. Moreover, we hypothesize the reversed reactions could have replaced canonical pathways for producing acetyl-CoA, essential for phospholipid biosynthesis. Additionally, we detected phylogenetically unrelated mevalonate pathway genes in metagenome-derived Rickettsiales sequences, likely indicating evolutionary convergent effects of independent horizontal gene transfer events. Accordingly, convergence, involving both gene acquisitions and losses, is highlighted as a relevant evolutionary phenomenon in Rickettsiales, possibly favoured by plasticity and comparable lifestyles, representing a potentially hidden origin of other more nuanced similarities among sub-lineages.Peer reviewe

The Genome Sequence of "Candidatus Fokinia solitaria": Insights on Reductive Evolution in Rickettsiales

"Candidatus Fokinia solitaria" is an obligate intracellular endosymbiont of a unicellular eukaryote, a ciliate of the genus Paramecium. Here, we present the genome sequence of this bacterium and subsequent analysis. Phylogenomic analysis confirmed the previously reported positioning of the symbiont within the "Candidatus Midichloriaceae" family (order Rickettsiales), as well as its high sequence divergence from other members of the family, indicative of fast sequence evolution. Consistently with this high evolutionary rate, a comparative genomic analysis revealed that the genome of this symbiont is the smallest of the Rickettsiales to date. The reduced genome does not present flagellar genes, nor the pathway for the biosynthesis of lipopolysaccharides (present in all the other so far sequenced members of the family "Candidatus Midichloriaceae") or genes for the Krebs cycle (present, although not always complete, in Rickettsiales). These results indicate an evolutionary trend toward a stronger dependence on the host, in comparison with other members of the family. Two alternative scenarios are compatible with our results; "Candidatus Fokinia solitaria" could be either a recently evolved, vertically transmitted mutualist, or a parasite with a high host-specificity

Comparative genomics of closely related strains of Klebsiella pneumoniae reveals genes possibly involved in colistin resistance

Strains of colistin-resistant Klebsiella pneumoniae are emerging worldwide, due to the increased use of this molecule in antibiotic-resistant nosocomial infections. Comparative genomics was performed on three closely related K. pneumoniae strains isolated from three patients in a single hospital in Bologna, Italy. Two of these isolates are colistin-resistant, while the third is sensitive to this antibiotic. The designed bioinformatic approach detected, among the three analyzed genomes, single nucleotide polymorphisms, insertions and deletions, specific patterns of gene presence and absence, in a total of 270 genes. These genes were analyzed by automatic and manual methods, to identify those potentially involved in colistin resistance, based on the data available in the literature and on the mechanism of action of colistin, the alteration of the outer membrane. Three of the identified genes (waaL, rfbA, vacJ), all presenting non-synonymous substitutions in the colistin resistant strains, resulted to be of special interest, due to the specific function of their protein products, involved in the biosynthesis of the outer bacterial membrane

The COVID-19 Pandemic Sparked off a Large-Scale Outbreak of Carbapenem-Resistant Acinetobacter baumannii from the Endemic Strains at an Italian Hospital

Acinetobacter baumannii is a nosocomial pathogen that poses a serious threat due to the rise of incidence of multidrug-resistant (MDR) strains. During the COVID-19 pandemic, MDR A. baumannii clones have caused several outbreaks worldwide. Here, we describe a detailed investigation of an MDR A. baumannii outbreak that occurred at Policlinico San Matteo (Pavia, Italy). A total of 96 A. baumannii strains, isolated between January and July 2020 from 41 inpatients (both SARS-CoV-2 positive and negative) in different wards, were characterized by phenotypic and genomic analyses combining Illumina and Nanopore sequencing. Antibiotic susceptibility testing revealed that all isolates were resistant to carbapenems, and the sequence analysis attributed this to the carbapenemase gene blaOXA-23. Virulence factor screening unveiled that all strains carried determinants for biofilm formation, while plasmid analysis revealed the presence of two plasmids, one of which was ~100 kbp long and encoded a phage sequence. A core genome-based phylogeny was inferred to integrate outbreak strain genomes with background genomes from public databases and the local surveillance program. All strains belonged to the globally disseminated sequence type 2 (ST2) clone and were mainly divided into two clades. Isolates from the outbreak clustered with surveillance isolates from 2019, suggesting that the outbreak was caused by two strains that were already circulating in the hospital before the start of the pandemic. The intensive spread of A. baumannii in the hospital was enhanced by the extreme emergency situation of the first COVID-19 pandemic wave that resulted in reduced attention to infection prevention and control practices. IMPORTANCE: The COVID-19 pandemic, especially during the first wave, posed a great challenge to the hospital management and generally promoted nosocomial pathogen dissemination. MDR A. baumannii can easily spread and persist for a long time on surfaces, causing outbreaks in health care settings. Infection prevention and control practices, epidemiological surveillance, and microbiological screening are fundamental in order to control such outbreaks. Here, we sequenced the genomes of 96 isolates from an outbreak of MDR A. baumannii strains using both short- and long-read technology in order to reconstruct the outbreak events in fine detail. The sequence data demonstrated that two endemic clones of MDR A. baumannii were the source of this large hospital outbreak during the first COVID-19 pandemic wave, confirming the effect of COVID-19 emergency disrupting the protection provided by the use of the standard prevention procedures

The choreography of the chemical defensome response to insecticide stress: insights into the Anopheles stephensi transcriptome using RNA-Seq

Animals respond to chemical stress with an array of gene families and pathways termed "chemical defensome". In arthropods, despite many defensome genes have been detected, how their activation is arranged during toxic exposure remains poorly understood. Here, we sequenced the transcriptome of Anopheles stephensi larvae exposed for six, 24 and 48 hours to the LD50 dose of the insecticide permethrin to monitor transcriptional changes of defensome genes across time. A total of 177 genes involved in insecticide defense were differentially expressed (DE) in at least one time-point, including genes encoding for Phase 0, I, II, III and antioxidant enzymes and for Heat Shock and Cuticular Proteins. Three major patterns emerged throughout time. First, most of DE genes were down-regulated at all time-points, suggesting a reallocation of energetic resources during insecticide stress. Second, single genes and clusters of genes turn off and on from six to 48 hours of treatment, showing a modulated response across time. Third, the number of up-regulated genes peaked at six hours and then decreased during exposure. Our results give a first picture of how defensome gene families respond against toxicants and provide a valuable resource for understanding how defensome genes work together during insecticide stress

Temporal dynamics of the ABC transporter response to insecticide treatment: insights from the malaria vector Anopheles stephensi

In insects, ABC transporters have been shown to contribute to defence/resistance to insecticides by reducing toxic concentrations in cells/tissues. Despite the extensive studies about this detoxifying mechanism, the temporal patterns of ABC transporter activation have been poorly investigated. Using the malaria vector Anopheles stephensi as a study system, we investigated the expression profile of ABC genes belonging to different subfamilies in permethrin-treated larvae at different time points (30 min to 48 h). Our results showed that the expression of ABCB and ABCG subfamily genes was upregulated at 1 h after treatment, with the highest expression observed at 6 h. Therefore, future investigations on the temporal dynamics of ABCgene expression will allow a better implementation of insecticide treatment regimens, including the use of specific inhibitors of ABC efflux pumps

Presence of wolbachia in three hymenopteran species : diprion pini (Hymenoptera: Diprionidae), neodiprion sertifer (Hymenoptera: Diprionidae), and dahlbominus fuscipennis (Hymenoptera: Eulophidae)

Sawflies are important pests of various plant species. Diprion pini (L.) and Neodiprion sertifer (Geoffroy) (Hymenoptera: Diprionidae) are two of the most important sawfly pests in Italy, and both species are parasitized by the hymenopteran parasitoid Dahlbominus fuscipennis (Zetterstedt). Bacterial endosymbionts are currently studied for their high potential in strategies of biocontrol in a number of insect species. In this study, we investigated the presence of symbiotic bacteria (Wolbachia and Cardinium) in the three species of hymenoptera mentioned earlier, both in wild and laboratory populations. Although all samples were negative for the presence of Cardinium, 100% prevalence for Wolbachia was detected, as all examined individuals resulted to be PCR positive. Furthermore, 16S rDNA and ftsZ gene sequencing indicated that all individuals from the three hymenopteran species are infected by a single Wolbachia strain. Additionally, we report the presence of gynandromorphic individuals in D. pini, both in wild and laboratory-reared populations. Heat treatments on D. pini colonies removed the Wolbachia symbionts, but they also prevented the development of adults

Scalable software framework for real-time data processing in the railway environment

Background: Ticks are obligate haematophagous ectoparasites of vertebrates and frequently parasitize avian species that can carry them across continents during their long-distance migrations. Ticks may have detrimental effects on the health state of their avian hosts, which can be either directly caused by blood-draining or mediated by microbial pathogens transmitted during the blood meal. Indeed, ticks host complex microbial communities, including bacterial pathogens and symbionts. Midichloria bacteria (Rickettsiales) are widespread tick endosymbionts that can be transmitted to vertebrate hosts during the tick bite, inducing an antibody response. Their actual role as infectious/pathogenic agents is, however, unclear. Methods: We screened for Midichloria DNA African ticks and blood samples collected from trans-Saharan migratory songbirds at their arrival in Europe during spring migration. Results: Tick infestation rate was 5.7%, with most ticks belonging to the Hyalomma marginatum species complex. Over 90% of Hyalomma ticks harboured DNA of Midichloria bacteria belonging to the monophylum associated with ticks. Midichloria DNA was detected in 43% of blood samples of avian hosts. Tick-infested adult birds were significantly more likely to test positive to the presence of Midichloria DNA than non-infested adults and second-year individuals, suggesting a long-term persistence of these bacteria within avian hosts. Tick parasitism was associated with a significantly delayed timing of spring migration of avian hosts but had no significant effects on body condition, whereas blood Midichloria DNA presence negatively affected fat deposits of tick-infested avian hosts. Conclusions: Our results show that ticks effectively transfer Midichloria bacteria to avian hosts, supporting the hypothesis that they are infectious to vertebrates. Bird infection likely enhances the horizontal spread of these bacteria across haematophagous ectoparasite populations. Moreover, we showed that Midichloria and tick parasitism have detrimental non-independent effects on avian host health during migration, highlighting the complexity of interactions involving ticks, their vertebrate hosts, and tick-borne bacteria

Supergroup C Wolbachia, mutualist symbionts of filarial nematodes, have a distinct genome structure

Wolbachia pipientis is possibly the most widespread endosymbiont of arthropods and nematodes. While all Wolbachia strains have historically been defined as a single species, 16 monophyletic clusters of diversity (called supergroups) have been described. Different supergroups have distinct host ranges and symbiotic relationships, ranging from mutualism to reproductive manipulation. In filarial nematodes, which include parasites responsible for major diseases of humans (such as Onchocerca volvulus, agent of river blindness) and companion animals (Dirofilaria immitis, the dog heartworm), Wolbachia has an obligate mutualist role and is the target of new treatment regimens. Here, we compare the genomes of eight Wolbachia strains, spanning the diversity of the major supergroups (A-F), analysing synteny, transposable element content, GC skew and gene loss or gain. We detected genomic features that differ between Wolbachia supergroups, most notably in the C and D clades from filarial nematodes. In particular, strains from supergroup C (symbionts of O. volvulus and D. immitis) present a pattern of GC skew, conserved synteny and lack of transposable elements, unique in the Wolbachia genus. These features could be the consequence of a distinct symbiotic relationship between C Wolbachia strains and their hosts, highlighting underappreciated differences between the mutualistic supergroups found within filarial nematodes