The best of both worlds: a proposal for further integration of Candidatus names into the International Code of Nomenclature of Prokaryotes.

The naming of prokaryotes is governed by the International Code of Nomenclature of Prokaryotes (ICNP) and partially by the International Code of Nomenclature for Algae, Fungi and Plants (ICN). Such codes must be able to determine names of taxa in a universal and unambiguous manner, thus serving as a common language across different fields and activities. This unity is undermined when a new code of nomenclature emerges that overlaps in scope with an established, time-tested code and uses the same format of names but assigns different nomenclatural status values to the names. The resulting nomenclatural confusion is not beneficial to the wider scientific community. Such ambiguity is expected to result from the establishment of the 'Code of Nomenclature of Prokaryotes Described from DNA Sequence Data' ('SeqCode'), which is in general and specific conflict with the ICNP and the ICN. Shortcomings in the interpretation of the ICNP may have exacerbated the incompatibility between the codes. It is reiterated as to why proposals to accept sequences as nomenclatural types of species and subspecies with validly published names, now implemented in the SeqCode, have not been implemented by the International Committee on Systematics of Prokaryotes (ICSP), which oversees the ICNP. The absence of certain regulations from the ICNP for the naming of as yet uncultivated prokaryotes is an acceptable scientific argument, although it does not justify the establishment of a separate code. Moreover, the proposals rejected by the ICSP are unnecessary to adequately regulate the naming of uncultivated prokaryotes. To provide a better service to the wider scientific community, an alternative proposal to emend the ICNP is presented, which would result in Candidatus names being regulated analogously to validly published names. This proposal is fully consistent with previous ICSP decisions, preserves the essential unity of nomenclature and avoids the expected nomenclatural confusion.

Proposal for Acinetobacter higginsii sp. nov. to accommodate organisms of human clinical origin previously classified as Acinetobacter genomic species 16.

In 1989, Bouvet and Jeanjean delineated five proteolytic genomic species (GS) of Acinetobacter, each with two to four human isolates. Three were later validly named, whereas the remaining two (GS15 and GS16) have been awaiting nomenclatural clarification. Here we present the results of the genus-wide taxonomic study of 13 human strains classified as GS16 (n=10) or GS15 (n=3). Based on core genome phylogenetic analysis, the strains formed two respective but closely related phylogroups within the Acinetobacter haemolytic clade. The intraspecies genomic average nucleotide identity based on blast (ANIb) values for GS16 and GS15 reached ≥94.9 % and ≥98.7, respectively, whereas ANIb values between them were 92.5-93.5% and those between them and the known species were ≤91.5 %. GS16 and GS15 could be differentiated from the other Acinetobacter species by their ability to lyse gelatin and sheep blood and to assimilate d,l-lactate, along with their inability to acidify d-glucose and assimilate glutarate. In contrast, GS16 and GS15 were indistinguishable from one another by metabolic/physiological features or whole-cell MALDI-TOF mass spectra. All the GS15/GS16 genomes contained genes encoding a class D ß-lactamase, Acinetobacter-derived cephalosporinase and aminoglycoside 6'-N-acetyltransferase. Searching NCBI databases revealed genome sequences of three additional isolates of GS16, but none of GS15. We conclude that our data support GS16 as representing a novel species, but leave the question of the taxonomic status of GS15 open, given its close relatedness to GS16 and the small number of available strains. We propose the name Acinetobacter higginsii sp. nov. for GS16, with the type strain NIPH 1872T (CCM 9243T=CIP 70.18T=ATCC 17988T).

Strain "Acinetobacter mesopotamicus" GC2 Does Not Represent a Novel Species, but Belongs to the Species Acinetobacter lwoffii as Revealed by Whole-Genome Sequence-Based Analysis.

The formal scientific name "Acinetobacter mesopotamicus" has recently been proposed for bacterial strain GC2 isolated from soil (Curr Microbiol 77:3192-3200). However, the recalculation of the average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) parameters revealed that their values for the genome sequences of GC2 and Acinetobacter lwoffii NCTC 5866T were as high as 95.7% and 66.1%, respectively. Moreover, the ANI and dDDH values for the genome of GC2 tested against those of 10 reference strains of A. lwoffii were 95.5-96.7% and 65.5-74.2%, respectively, and fall within the intraspecies range of these values known for A. lwoffii. These data indicate that strain GC2 belongs to A. lwoffii and contradicts the results of the authors of the proposal, who found the ANI and dDDH values for the GC2 and A. lwoffii DSM 2403T genomes to be 88.87% and 36.1%, respectively. The misclassification of strain GC2 is likely to result from the use of an incorrect reference genome sequence for in silico comparative analyses.

Differentiation of Taxonomically Closely Related Species of the Genus Acinetobacter Using Raman Spectroscopy and Chemometrics.

In recent years, several efforts have been made to develop quick and low cost bacterial identification methods. Genotypic methods, despite their accuracy, are laborious and time consuming, leaving spectroscopic methods as a potential alternative. Mass and infrared spectroscopy are among the most reconnoitered techniques for this purpose, with Raman having been practically unexplored. Some species of the bacterial genus Acinetobacter are recognized as etiological agents of nosocomial infections associated with high rates of mortality and morbidity, which makes their accurate identification important. The goal of this study was to assess the ability of Raman spectroscopy to discriminate between 16 Acinetobacter species belonging to two phylogroups containing taxonomically closely related species, that is, the Acinetobacter baumannii-Acinetobacter calcoaceticus complex (six species) and haemolytic clade (10 species). Bacterial spectra were acquired without the need for any sample pre-treatment and were further analyzed with multivariate data analysis, namely partial least squares discriminant analysis (PLSDA). Species discrimination was achieved through a series of sequential PLSDA models, with the percentage of correct species assignments ranging from 72.1% to 98.7%. The obtained results suggest that Raman spectroscopy is a promising alternative for identification of Acinetobacter species.

Acinetobacter wuhouensis sp. nov., isolated from hospital sewage.

We recovered eight strains of the genus Acinetobacter from hospital sewage at West China Hospital in Chengdu, China. Based on the comparative analysis of the rpoB sequence, these strains formed a strongly supported and internally coherent cluster (intra-cluster identity of ≥98.0 %), which was clearly separated from all known Acinetobacter species (≤91.1 %). The eight strains also formed a tight and distinct cluster based on the genus-wide comparison of whole-cell mass fingerprints generated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. In addition, the combination of their ability to assimilate 2,3-butanediol and phenylacetate, but not 4-hydroxybenzoate, and the inability to grow at 37 °C could distinguish these eight strains from all known Acinetobacter species. Whole-genomic sequencing has been performed for two selected strains, WCHA60T and WCHA62. There were 96.65 % average nucleotide identity (ANI) and 72 % in silico DNA-DNA hybridization (isDDH) values between WCHA60T and WCHA62, suggesting that the two strains indeed belonged to the same species. In contrast, the ANI and isDDH values between the two strains and the known Acinetobacter species were <83 and <30 %, respectively; both of which were far below the cut-off to define a bacterial species. Therefore, the eight strains should be considered to represent a novel species of the genus Acinetobacter, for which the name Acinetobacterwuhouensis sp. nov. is proposed. The type strain is WCHA60T (=CCTCC AB 2016204T=GDMCC 1.1100T=KCTC 52505T).

Ability of phages to infect Acinetobacter calcoaceticus-Acinetobacter baumannii complex species through acquisition of different pectate lyase depolymerase domains.

Bacteriophages are ubiquitous in nature and represent a vast repository of genetic diversity, which is driven by the endless coevolution cycle with a diversified group of bacterial hosts. Studying phage-host interactions is important to gain novel insights into their dynamic adaptation. In this study, we isolated 12 phages infecting species of the Acinetobacter baumannii-Acinetobacter calcoaceticus complex which exhibited a narrow host range and similar morphological features (podoviruses with short tails of 9-12 nm and isometric heads of 50-60 nm). Notably, the alignment of the newly sequenced phage genomes (40-41 kb of DNA length) and all Acinetobacter podoviruses deposited in Genbank has shown high synteny, regardless of the date and source of isolation that spans from America to Europe and Asia. Interestingly, the C-terminal pectate lyase domain of these phage tail fibres is often the only difference found among these viral genomes, demonstrating a very specific genomic variation during the course of their evolution. We proved that the pectate lyase domain is responsible for phage depolymerase activity and binding to specific Acinetobacter bacterial capsules. We discuss how this mechanism of phage-host co-evolution impacts the tail specificity apparatus of Acinetobacter podoviruses.

Acinetobacter guangdongensis Feng et al. 2014 is a junior heterotypic synonym of Acinetobacter indicus Malhotra et al. 2012.

A draft whole-genome sequence was obtained for Acinetobacter guangdongensis strain KCTC 42012T and compared against those of the type strains of all Acinetobacter species with validly published names. High similarity was found to Acinetobacter indicus CCM 7832T (average nucleotide identity based on blast and digital DNA-DNA hybridization values of 96.3 and 70.4 %, respectively). In addition, the metabolic, physiological and chemotaxonomic features of KCTC 42012T were shown to be congruent with those of A. indicus. We conclude that Acinetobacter guangdongensisFeng et al. 2014 is a later heterotypic synonym of Acinetobacter indicus Malhotra et al. 2012.

Acinetobacter colistiniresistens sp. nov. (formerly genomic species 13 sensu Bouvet and Jeanjean and genomic species 14 sensu Tjernberg and Ursing), isolated from human infections and characterized by intrinsic resistance to polymyxins.

Strains of the genusAcinetobacter, classified as genomic species 13BJ/14TU have been previously associated with human infections and resistance to colistin. To clarify the taxonomy of this provisional group, we investigated 24 strains that have been isolated from humans since the 1960s in 10 countries. The genus-wide analysis of the rpoB and gyrB sequences of all strains and whole-genome sequences of strains representing different rpoB/gyrB genotypes showed that the 24 strains formed a distinct monophyletic group within the so-called haemolytic clade of the genus Acinetobacter. The distinctness of the group at the species level was supported by the results of the cluster analysis of the whole-cell protein fingerprints generated by matrix-assisted laser desorption ionization-time-of-flight MS. The 24 strains had very similar metabolic features and could be distinguished from other members of the genus by the combination of strong haemolytic and proteolytic activities and the ability to oxidize d-glucose and grow on phenylacetate and/or l-phenylalanine. The minimum inhibitory concentrations of the 24 strains to colistin and polymyxin B ranged from 16 to 64 mgl-1 and from 4 to 32 mgl-1, respectively, so uniformly reaching the current clinical resistance breakpoint (4 mg l-1) for these drugs. Genus-wide comparison revealed that such a consistently high level of resistance to polymyxins is a unique feature among species of the genus Acinetobacter,which occur in humans. We conclude that genomic species 13BJ/14TU represents a biologically meaningful and medically relevant species, for which the name Acinetobacter colistiniresistens sp. nov. is proposed. The type strain is NIPH 2036T (=CCM 8641T=CIP 110478T=CCUG 67966T=CNCTC 7573T).

Origin in Acinetobacter gyllenbergii and dissemination of aminoglycoside-modifying enzyme AAC(6')-Ih.

OBJECTIVES: The aac(6')-Ih gene encoding aminoglycoside 6'-N-acetyltransferase type I subtype h [AAC(6')-Ih] is plasmid-borne in Acinetobacter baumannii where it confers high-level amikacin resistance, but its origin remains unknown. We searched for the gene in the genomes of a collection of 133 Acinetobacter spp. and studied its species specificity, expression and dissemination. METHODS: Gene copy number was determined by quantitative PCR, expression by quantitative RT-PCR, MIC by microdilution and transfer by plasmid mobilization. RESULTS: The aac(6')-Ih gene was present in the chromosome of the two Acinetobacter gyllenbergii of the collection and was detected in all seven A. gyllenbergii clinical isolates. They had indistinguishable flanking regions indicating that the gene was intrinsic to this species. A. baumannii PIS Aba23 promoters were provided by insertion of ISAba23, which disrupted the Pnative promoter in A. gyllenbergii. Both types of promoters were similarly potent in Escherichia coli and A. baumannii. Aminoglycoside MICs for A. baumannii harbouring pIP1858 were higher than for A. gyllenbergii due to gene dosage. The non-self-transferable plasmid could be mobilized to other A. baumannii cells by the broad host range plasmid RP4. CONCLUSIONS: We have found the origin of aac(6')-Ih in A. gyllenbergii, a species isolated, although rarely, in humans, and documented that dissemination of this gene is restricted to the Acinetobacter genus.

Acinetobacter pakistanensis Abbas et al. 2014 is a later heterotypic synonym of Acinetobacter bohemicus Krizova et al. 2014.

Two novel species names, Acinetobacter bohemicus and Acinetobacter pakistanensis, appeared on validation list no. 161 (January 2015) under priority numbers 26 and 28, respectively. As the published data suggested a high similarity of the organisms associated with these names, we aimed to define their taxonomic relationship. The study set included all strains used in the original nomenclatural proposals, i.e. 25 strains of A. bohemicus and one strain of A. pakistanensis. The average nucleotide identity values (95.9 and 96.1 % based on blast and MUMmer, respectively) between the whole-genome sequences of A. bohemicus ANC 3994T and A. pakistanensis KCTC 42081T supported the identity of these type strains at the species level. Based on the genus-wide comparative analyses of the rpoB sequences and whole-cell fingerprints generated by matrix-assisted laser desorption/ionization-time-of-flight MS, A. pakistanensis KCTC 42081T fell within the respective clusters formed by the 25 A. bohemicus strains. The same picture was obtained on the basis of comparative analysis of 16S rRNA gene sequences of KCTC 42081T and three A. bohemicus strains. Finally, the metabolic and physiological features of KCTC 42081T were found to be congruent with those of A. bohemicus. Based on these results, we conclude that Acinetobacter pakistanensis is a later heterotypic synonym of Acinetobacter bohemicus.

Acinetobactercelticus sp. nov., a psychrotolerant species widespread in natural soil and water ecosystems.

A novel, taxonomically unique group of six strains of the genus Acinetobacter was discovered during an exploratory study on strains culturable from soil and water natural ecosystems in the Bohemian part of the Czech Republic. Based on the comparative analyses of the 16S rRNA gene, gyrB and rpoB sequences, these strains formed strongly supported and internally coherent clusters (intracluster identities of ≥99.9, ≥96.1 and ≥97.3 %, respectively), which were clearly separated from all known species of the genus Acinetobacter (≤98.7, ≤83.2 and ≤88.9 %, respectively). The distinctness of the group at the species level was evidenced also by the results of the genus-wide analyses of the whole-cell mass fingerprints of the six strains generated by matrix-assisted laser desorption/ionization-time-of-flight MS and the whole-genome sequence of a group member, ANC 4603T. Compared with the known species of the genus Acinetobacter, all six strains exhibited a unique phenotype, characterized by psychrotolerance (growth at 1 °C through 28 °C), the inability to grow at 32 °C and the ability to assimilate l-aspartate and malonate but not 2,3-butanediol or citrate. Based on these results, the name Acinetobacter celticus sp. nov. is proposed for the taxon represented by the six strains. The type strain is ANC 4603T (=CCM 8700T=CCUG 69239T=CNCTC 7549T).

Acinetobacter pragensis sp. nov., found in soil and water ecosystems.

This study aimed to define the taxonomic status of a novel, phenetically distinct group of seven strains belonging to the genus Acinetobacter, which were isolated from environmental soil and water samples collected in Central Bohemia, Czech Republic. Comparative sequence analyses of the 16S rRNA, gyrB and rpoB genes showed that all these strains formed respective tight clusters (intracluster sequence similarities of ≥99.8, ≥98.1 and ≥98.3 %, respectively), which were distant from all known Acinetobacter species (≤98.2, ≤84.0 and ≤88.9 %, respectively). The average nucleotide identity and digital DNA-DNA hybridization values (≤83.5 and ≤27.4 %, respectively) between the whole-genome sequence of a group representative (strain ANC 4149T) and those of known taxa were far below the thresholds used to discriminate between bacterial species. The seven strains also formed a tight and distinct cluster based on the genus-wide comparison of whole-cell mass fingerprints generated by matrix-assisted laser desorption/ionization time-of-flight MS and could be distinguished from all other members of the genus Acinetobacter by the combination of their ability to assimilate glutarate and l-tartrate and inability to grow at 37 °C and on l-aspartate. It is concluded that the seven strains represent a novel species for which the name Acinetobacter pragensis sp. nov. is proposed. The type strain is ANC 4149T (=CCM 8637T=CCUG 67962T=CNCTC 7530T).

Taxonomy of haemolytic and/or proteolytic strains of the genus Acinetobacter with the proposal of Acinetobacter courvalinii sp. nov. (genomic species 14 sensu Bouvet & Jeanjean), Acinetobacter dispersus sp. nov. (genomic species 17), Acinetobacter modestus sp. nov., Acinetobacter proteolyticus sp. nov. and Acinetobacter vivianii sp. nov.

We aimed to define the taxonomic status of 40 haemolytic and/or proteolytic strains of the genus Acinetobacter which were previously classified into five putative species termed as genomic species 14BJ (n=9), genomic species 17 (n=9), taxon 18 (n=7), taxon 19 (n=6) and taxon 20 (n=9). The strains were recovered mostly from human clinical specimens or soil and water ecosystems and were highly diverse in geographical origin and time of isolation. Comparative analysis of the rpoB and gyrB gene sequences of all strains, and the whole-genome sequences of selected strains, showed that these putative species formed five respective, well-supported clusters within a distinct clade of the genus Acinetobacter which typically, although not exclusively, encompasses strains with strong haemolytic activity. The whole-genome-based average nucleotide identity (ANIb) values supported the species status of each of these clusters. Moreover, the distinctness and coherence of the clusters were supported by whole-cell profiling based on MALDI-TOF MS. Congruent with these findings were the results of metabolic and physiological testing. We conclude that the five putative taxa represent respective novel species, for which the names Acinetobacter courvalinii sp. nov. (type strain ANC 3623T=CCUG 67960T=CIP 110480T=CCM 8635T), Acinetobacter dispersus sp. nov. (type strain ANC 4105T=CCUG 67961T=CIP 110500T=CCM 8636T), Acinetobacter modestus sp. nov. (type strain NIPH 236T=CCUG 67964T=CIP 110444T=CCM 8639T), Acinetobacter proteolyticus sp. nov. (type strain NIPH 809T=CCUG 67965T=CIP 110482T = CCM 8640T) and Acinetobacter vivianii sp. nov. (type strain NIPH 2168T=CCUG 67967T=CIP 110483T=CCM 8642T) are proposed.

A taxonomically unique Acinetobacter strain with proteolytic and hemolytic activities recovered from a patient with a soft tissue injury.

A taxonomically unique bacterial strain, Acinetobacter sp. A47, has been recovered from several soft tissue samples from a patient undergoing reconstructive surgery owing to a traumatic amputation. The results of 16S rRNA, rpoB, and gyrB gene comparative sequence analyses showed that A47 does not belong to any of the hitherto-known taxa and may represent an as-yet-unknown Acinetobacter species. The recognition of this novel organism contributes to our knowledge of the taxonomic complexity underlying infections caused by Acinetobacter.

Acinetobacter albensis sp. nov., isolated from natural soil and water ecosystems.

We have studied the taxonomic position of a phenetically unique group of eight strains of the genus Acinetobacter which were isolated from soil and water samples collected in protected landscape areas in the Czech Republic. Each of the comparative sequence analyses of the 16S rRNA, gyrB and rpoB genes showed that the eight strains formed a cohesive and tight cluster (intracluster sequence identities of ≥ 99.9 %, ≥ 98.5 % and ≥ 97.7 %, respectively), which was clearly separated from all hitherto known species of the genus Acinetobacter ( ≤ 98.6 %, ≤ 84.5 % and ≤ 89.3 %, respectively). Congruent with these findings were the results of comparative sequence analysis of three additional housekeeping genes (gltA, pyrG and recA). This genotypic distinctness was mirrored by the uniqueness of the combination of a number of independent phenotypic markers including the whole-cell spectra produced by matrix-assisted laser desorption ionization time-of-flight (MALDI-ToF) MS and physiological and metabolic features. The most useful phenotypic features to differentiate the eight strains from all known species of the genus Acinetobacter were the ability to assimilate tricarballylate and the inability to grow at 35 °C or to assimilate ethanol or l-histidine. We conclude that the eight strains represent a novel environmental species for which the name Acinetobacter albensis sp. nov. is proposed. The type strain is ANC 4874T ( = CCUG 67281T = CCM 8611T).

Acinetobacter seifertii sp. nov., a member of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex isolated from human clinical specimens.

This study aimed to define the taxonomic status of a phenetically distinct group of 16 strains that corresponds to Acinetobacter genomic species 'close to 13TU', a provisional genomic species of the Acinetobacter calcoaceticus-Acinetobacter baumannii (ACB) complex recognized by Gerner-Smidt and Tjernberg in 1993. These strains have been isolated in different countries since the early 1990s and were mostly recovered from human clinical specimens. They were compared with 45 reference strains representing the known taxa of the ACB complex using taxonomic methods relevant to the genus Acinetobacter. Based on sequence analysis of the concatenated partial sequences (2976 bp) of seven housekeeping genes, the 16 strains formed a tight and well-supported cluster (intracluster sequence identity of ≥98.4 %) that was clearly separated from the other members of the ACB complex (≤94.7 %). The species status of the group was supported by average nucleotide identity values of ≤91.7 % between the whole genome sequence of representative strain NIPH 973(T) (NCBI accession no. APOO00000000) and those of the other species. In addition, whole-cell matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) MS analyses indicated the distinctness of the group at the protein level. Metabolic and physiological tests revealed several typical features of the group, although they did not allow its reliable differentiation from the other members of the ACB complex. We conclude that the 16 strains represent a distinct novel species, for which we propose the name Acinetobacter seifertii sp. nov. The type strain is NIPH 973(T) ( = CIP 110471(T) = CCUG 34785(T) = CCM 8535(T)).