1. |
Pidiyar VJ,
Jangid K,
Dayananda KM,
Kaznowski A,
Gonzalez JM,
Patole MS,
Shouche YS,
( 2003 ) Phylogenetic affiliation of Aeromonas culicicola MTCC 3249(T) based on gyrB gene sequence and PCR-amplicon sequence analysis of cytolytic enterotoxin gene. PMID : 12866846 : DOI : 10.1078/072320203322346047 Abstract >>
We determined the gyrB gene sequences of all 17 hybridizations groups of Aeromonas. Phylogenetic trees showing the evolutionary relatedness of gyrB and 16S rRNA genes in the type strains of Aeromonas were compared. Using this approach, we determined the phylogenetic position of Aeromonas culicicola MTCC 3249(T), isolated from midgut of Culex quinquefasciatus. In the gyrB based-analysis A. culicicola MTCC 3249(T) grouped with A. veronii whereas, it grouped with A. jandaei in the 16S rRNA based tree. The number of nucleotide differences in 16S rRNA sequences was less than found with the gyrB sequence data. Most of the observed nucleotide differences in the gyrB gene were synonymous. The Cophenetic Correlation Coefficient (CCC) for gyrB sequences was 0.87 indicating this gene to be a better molecular chronometer compared to 16S rRNA for delineation of Aeromonas species. This strain was found to be positive for the cytolytic enterotoxin gene. PCR-Amplicon Sequence Analysis (PCR-ASA) of this gene showed that the isolate is affiliated to type I and is potentially pathogenic. These PCR-ASA results agreed in part with the gyrB sequence results.
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2. |
Yáñez MA,
Catalán V,
Apráiz D,
Figueras MJ,
Martínez-Murcia AJ,
( 2003 ) Phylogenetic analysis of members of the genus Aeromonas based on gyrB gene sequences. PMID : 12807216 : DOI : 10.1099/ijs.0.02443-0 Abstract >>
The phylogenetic relationships of all known species of the genus Aeromonas were investigated by using the sequence of gyrB, a gene that encodes the B-subunit of DNA gyrase. Nucleotide sequences of gyrB were determined from 53 Aeromonas strains, including some new isolates, which were also characterized by analysis of the 16S rDNA variable regions. The results support the recognition of the family Aeromonadaceae, as distinct from Plesiomonas shigelloides and other enteric bacteria. This phylogenetic marker revealed strain groupings that are consistent with the taxonomic organization of all Aeromonas species described to date. In particular, gyrB results agreed with 16S rDNA analysis; moreover, the former showed a higher capacity to differentiate between species. The present analysis was useful for the elucidation of reported discrepancies between different DNA-DNA hybridization sets. Additionally, due to the sequence diversity found at the intraspecies level, gyrB is proposed as a useful target for simultaneous identification of species and strains. In conclusion, the gyrB gene has proved to be an excellent molecular chronometer for phylogenetic studies of the genus Aeromonas.
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3. |
Saavedra MJ,
Figueras MJ,
Martínez-Murcia AJ,
( 2006 ) Updated phylogeny of the genus Aeromonas. PMID : 17012583 : DOI : 10.1099/ijs.0.64351-0 Abstract >>
Recent phylogenetic studies of the genus Aeromonas based on gyrB and rpoD gene sequences have improved the phylogeny based on 16S rRNA gene sequences first published in 1992, particularly in the ability to split closely related species. These studies did not include the recently described species Aeromonas simiae and Aeromonas molluscorum and only a single strain of Aeromonas culicicola was available for analysis at that time. In the present work, these Aeromonas species and newly isolated strains of A. culicicola were examined. Sequence analysis indicates that A. simiae and A. molluscorum belong to non-described phylogenetic lines of descent within this genus, which supports the original description of both species. The most closely related species are Aeromonas schubertii and Aeromonas encheleia, respectively, which is consistent with 16S rRNA gene sequencing results. However, while the five strains of A. molluscorum showed nucleotide differences in their gyrB and rpoD gene sequences, the only two known A. simiae strains exhibited identical gene sequences, suggesting that they are isolates of the same strain. On the basis of the rpoD gene sequence phylogeny, A. culicicola strains from the original description and new isolates from drinking water and ornamental fish clustered within the species Aeromonas veronii, suggesting inconsistencies with previous results. Other strains with previously controversial taxonomy and new isolates from other studies were included in this study in order to clarify their phylogenetic affiliation at the species level.
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4. |
Sen K,
( 2005 ) Development of a rapid identification method for Aeromonas species by multiplex-PCR. PMID : 16333335 : DOI : 10.1139/w05-089 Abstract >>
Existing biochemical methods cannot distinguish among some species of Aeromonads, while genetic methods are labor intensive. In this study, primers were developed to three genes of Aeromonas: lipase, elastase, and DNA gyraseB. In addition, six previously described primer sets, five corresponding to species-specific signature regions of the 16S rRNA gene from A. veronii, A. popoffii, A. caviae, A. jandaei, and A. schubertii, respectively, and one corresponding to A. hydrophila specific lipase (hydrolipase), were chosen. The primer sets were combined in a series of multiplex-PCR (mPCR) assays against 38 previously characterized strains. Following PCR, each species was distinguished by the production of a unique combination of amplicons. When the assays were tested using 63 drinking water isolates, there was complete agreement in the species identification (ID) for 59 isolates, with ID established by biochemical assays. Sequencing the gyrB and the 16S rRNA gene from the remaining four strains established that the ID obtained by mPCR was correct for three strains. For only one strain, no consensus ID could be obtained. A rapid and reliable method for identification of different Aeromonas species is proposed that does not require restriction enzyme digestions, thus simplifying and speeding up the process.
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5. |
Soler L,
Yáñez MA,
Chacon MR,
Aguilera-Arreola MG,
Catalán V,
Figueras MJ,
Martínez-Murcia AJ,
( 2004 ) Phylogenetic analysis of the genus Aeromonas based on two housekeeping genes. PMID : 15388703 : DOI : 10.1099/ijs.0.03048-0 Abstract >>
The phylogenetic relationships of all known species of the genus Aeromonas, and especially Aeromonas bestiarum and Aeromonas salmonicida, were investigated on 70 strains using the rpoD sequence, which encodes the sigma70 factor. This analysis was complemented with the sequence of gyrB, which has already proven useful for determining the phylogenetic relationships in the genus. Nucleotide sequences of rpoD and gyrB showed that both genes had similar substitution rates (< 2 %) and a similar number of variable positions (34 % for rpoD versus 32 % for gyrB). Strain groupings by analysis of rpoD, gyrB and a combination of both genes were consistent with the taxonomic organization of all Aeromonas species described to date. However, the simultaneous analysis of both clocks improved the reliability and the power to differentiate, in particular, closely related taxa. At the inter-species level, gyrB showed a better resolution for differentiating Aeromonas sp. HG11/Aeromonas encheleia and Aeromonas veronii/Aeromonas culicicola/Aeromonas allosaccharophila, while rpoD more clearly differentiated A. salmonicida from A. bestiarum. The analysis of rpoD provided initial evidence for clear phylogenetic divergence between the latter two species.
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6. |
Figueira V,
Vaz-Moreira I,
Silva M,
Manaia CM,
( 2011 ) Diversity and antibiotic resistance of Aeromonas spp. in drinking and waste water treatment plants. PMID : 21907383 : DOI : 10.1016/j.watres.2011.08.021 Abstract >>
The taxonomic diversity and antibiotic resistance phenotypes of aeromonads were examined in samples from drinking and waste water treatment plants (surface, ground and disinfected water in a drinking water treatment plant, and raw and treated waste water) and tap water. Bacteria identification and intra-species variation were determined based on the analysis of the 16S rRNA, gyrB and cpn60 gene sequences. Resistance phenotypes were determined using the disc diffusion method. Aeromonas veronii prevailed in raw surface water, Aeromonas hydrophyla in ozonated water, and Aeromonas media and Aeromonas puntacta in waste water. No aeromonads were detected in ground water, after the chlorination tank or in tap water. Resistance to ceftazidime or meropenem was detected in isolates from the drinking water treatment plant and waste water isolates were intrinsically resistant to nalidixic acid. Most of the times, quinolone resistance was associated with the gyrA mutation in serine 83. The gene qnrS, but not the genes qnrA, B, C, D or qepA, was detected in both surface and waste water isolates. The gene aac(6')-ib-cr was detected in different waste water strains isolated in the presence of ciprofloxacin. Both quinolone resistance genes were detected only in the species A. media. This is the first study tracking antimicrobial resistance in aeromonads in drinking, tap and waste water and the importance of these bacteria as vectors of resistance in aquatic environments is discussed.
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7. |
Fontes MC,
Saavedra MJ,
Martins C,
Martínez-Murcia AJ,
( 2011 ) Phylogenetic identification of Aeromonas from pigs slaughtered for consumption in slaughterhouses at the North of Portugal. PMID : 21402427 : DOI : 10.1016/j.ijfoodmicro.2011.02.010 Abstract >>
In the present study, 710 isolates of Aeromonas spp. have been collected from pig carcasses, diaphragm muscle, faeces, dehairing equipment and water in slaughterhouses at the North of Portugal. The isolates were obtained from a total of 154 samples. All presumptive Aeromonas isolates were subjected to ERIC-PCR analysis and those which presented a different pattern were taken and the species classified by gyrB gene sequencing. We have found the species A. hydrophila, A. salmonicida, A. bestiarum, A. caviae, A. media, A. veronii, A. allosaccharophila, A. simiae and A. aquariorum. To our knowledge, this extent of Aeromonas species diversity has not been previously described from meat or from the slaughter environment, perhaps due to the unreliability of available identification methods. A noticeable level of isolate redundancy (strains with identical gyrB sequence) from different samples collected in different dates was also obtained, indicating that only a few predominant strains of these species persist at the slaughter system. It is also important to emphasise the presence of Aeromonas species previously associated with illness in man.
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8. |
Martinez-Murcia AJ,
Monera A,
Saavedra MJ,
Oncina R,
Lopez-Alvarez M,
Lara E,
Figueras MJ,
( 2011 ) Multilocus phylogenetic analysis of the genus Aeromonas. PMID : 21353754 : DOI : 10.1016/j.syapm.2010.11.014 Abstract >>
A broad multilocus phylogenetic analysis (MLPA) of the representative diversity of a genus offers the opportunity to incorporate concatenated inter-species phylogenies into bacterial systematics. Recent analyses based on single housekeeping genes have provided coherent phylogenies of Aeromonas. However, to date, a multi-gene phylogenetic analysis has never been tackled. In the present study, the intra- and inter-species phylogenetic relationships of 115 strains representing all Aeromonas species described to date were investigated by MLPA. The study included the independent analysis of seven single gene fragments (gyrB, rpoD, recA, dnaJ, gyrA, dnaX, and atpD), and the tree resulting from the concatenated 4705 bp sequence. The phylogenies obtained were consistent with each other, and clustering agreed with the Aeromonas taxonomy recognized to date. The highest clustering robustness was found for the concatenated tree (i.e. all Aeromonas species split into 100% bootstrap clusters). Both possible chronometric distortions and poor resolution encountered when using single-gene analysis were buffered in the concatenated MLPA tree. However, reliable phylogenetic species delineation required an MLPA including several "bona fide" strains representing all described species.
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9. |
Farfán M,
Miñana-Galbis D,
Garreta A,
Lorén JG,
Fusté MC,
( 2010 ) Malate dehydrogenase: a useful phylogenetic marker for the genus Aeromonas. PMID : 21095084 : DOI : 10.1016/j.syapm.2010.09.005 Abstract >>
The reconstruction of correct genealogies among biological entities, the estimation of the divergence time between organisms or the study of the different events that occur along evolutionary lineages are not always based on suitable genes. For reliable results, it is necessary to look at full-length sequences of genes under stabilizing selection (neutral or purifying) and behaving as good molecular clocks. In bacteria it has been proved that the malate dehydrogenase gene (mdh) can be used to determine the inter- and intraspecies divergence, and hence this gene constitutes a potential marker for phylogeny and bacterial population genetics. We have sequenced the full-length mdh gene in 36 type and reference strains of Aeromonas. The species grouping obtained in the phylogenetic tree derived from mdh sequences was in agreement with that currently accepted for the genus Aeromonas. The maximum likelihood models applied to our sequences indicated that the mdh gene is highly conserved among the Aeromonas species and the main evolutionary force acting on it is purifying selection. Only two sites under potential diversifying selection were identified (T 108 and S 193). In order to determine if these two residues could have an influence on the MDH structure, we mapped them in a three-dimensional model constructed from the sequence of A. hydrophila using the human mitochondrial MDH as a template. The presence of purifying selection together with the linear relationship between substitutions and gene divergence makes the mdh an excellent candidate gene for a phylogeny of Aeromonas and probably for other bacterial groups.
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10. |
Girlich D,
Poirel L,
Nordmann P,
( 2011 ) Diversity of clavulanic acid-inhibited extended-spectrum �]-lactamases in Aeromonas spp. from the Seine River, Paris, France. PMID : 21149627 : DOI : 10.1128/AAC.00921-10 PMC : PMC3067085 Abstract >>
Environmental Aeromonas sp. isolates resistant to ceftazidime were recovered during an environmental survey performed with water samples from the Seine River, in Paris, France, in November 2009. Selected isolates were identified by sequencing of the 16S rRNA and rpoB genes. PCR and cloning experiments were used to identify broad-spectrum-�]-lactamase-encoding genes and their genetic context. Clavulanic acid-inhibited extended-spectrum-�]-lactamase (ESBL) genes were identified in 71% of the Aeromonas sp. isolates. A variety of ESBL genes were detected, including bla(VEB-1a), bla(SHV-12), bla(PER-1), bla(PER-6), bla(TLA-2), and bla(GES-7), suggesting an aquatic reservoir of those ESBL genes. Moreover, the repeated elements and different insertion sequences were identified in association with the bla(PER-6) and the bla(VEB-1a) genes, respectively, indicating a wide diversity of mobilization events, making Aeromonas spp. a vehicle for ESBL dissemination.
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11. |
Lorén JG,
Farfán M,
Miñana-Galbis D,
Fusté MC,
( 2010 ) Prediction of whole-genome DNA G+C content within the genus Aeromonas based on housekeeping gene sequences. PMID : 20466501 : DOI : 10.1016/j.syapm.2010.03.007 Abstract >>
Different methods are available to determine the G+C content (e.g. thermal denaturation temperature or high performance liquid chromatography, HPLC), but obtained values may differ significantly between strains, as well as between laboratories. Recently, several authors have demonstrated that the genomic DNA G+C content of prokaryotes can be reliably estimated from one or several protein coding gene nucleotide sequences. Few G+C content values have been published for the Aeromonas species described and the data, when available, are often incomplete or provide only a range of values. Our aim in this current work was twofold. First, the genomic G+C content of the type or reference strains of all species and subspecies of the genus Aeromonas was determined with a traditional experimental method in the same laboratory. Second, we wanted to see if the sequence-based method to estimate the G+C content described by Fournier et al. [7] could be applied to determine the G+C content of the different species of Aeromonas from the sequences of the genes used in taxonomy or phylogeny for this genus.
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12. |
Wang W,
Wang L,
Shao Z,
( 2010 ) Diversity and abundance of oil-degrading bacteria and alkane hydroxylase (alkB) genes in the subtropical seawater of Xiamen Island. PMID : 20683589 : DOI : 10.1007/s00248-010-9724-4 Abstract >>
In this report, the diversity of oil-degrading bacteria and alkB gene was surveyed in the seawater around Xiamen Island. Forty-four isolates unique in 16S rRNA sequence were obtained after enrichment with crude oil. Most of the obtained isolates exhibited growth with diesel oil and crude oil. alkB genes were positively detected in 16 isolates by degenerate polymerase chain reaction (PCR). And for the first time, alkB genes were found in bacteria of Gallaecimonas, Castellaniella, Paracoccus, and Leucobacter. Additional 29 alkB sequences were retrieved from genomic DNA of the oil-degrading communities. Phylogenetic analysis showed that the obtained alkB genes formed five groups, most of which exhibited 60-80% similarity at the amino acid level with sequences retrieved from the GenBank database. Furthermore, the abundance of alkB genes in seawater was examined by real-time PCR. The results showed that alkB genes of each group in situ ranged from about 3 �� 10(3) to 3 �� 10(5) copies L(-1), with the homologs of Alcanivorax and Pseudomonas being the most predominant. Bacteria of Alcanivorax, Acinetobacter, and Pseudomonas are important oil degraders in this area; while those frequently reported in other area, like Oleiphilus spp., Oleispira spp., and Thalassolituus spp. were not found in our report. These results indicate that bacteria and genes involved in oil degradation are quite diverse, and may have restriction in geographic distribution in some species.
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13. |
Miñana-Galbis D,
Urbizu-Serrano A,
Farfán M,
Fusté MC,
Lorén JG,
( 2009 ) Phylogenetic analysis and identification of Aeromonas species based on sequencing of the cpn60 universal target. PMID : 19567585 : DOI : 10.1099/ijs.0.005413-0 Abstract >>
An analysis of the universal target (UT) sequence from the cpn60 gene was performed in order to evaluate its usefulness in phylogenetic and taxonomic studies and as an identification marker for the genus Aeromonas. Sequences of 555 bp, corresponding to the UT region, were obtained from a collection of 35 strains representing all of the species and subspecies of Aeromonas. From the analysis of these sequences, a range of divergence of 0-23.3% was obtained, with a mean of 11.2+/-0.9%. Comparative analyses between cpn60 and gyrB, rpoD and 16S rRNA gene sequences were carried out from the same Aeromonas strain collection. Sequences of the cpn60 UT region showed similar discriminatory power to gyrB and rpoD sequences. The phylogenetic relationships inferred from cpn60 sequence distances indicated an excellent correlation with the present affiliation of Aeromonas species with the exception of Aeromonas hydrophila subsp. dhakensis, which appeared in a separate phylogenetic line, and Aeromonas sharmana, which exhibited a very loose phylogenetic relationship to the genus Aeromonas. Sequencing of cpn60 from 33 additional Aeromonas strains also allowed us to establish intra- and interspecific threshold values. Intraspecific divergence rates were
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14. |
Alperi A,
Figueras MJ,
Inza I,
Martínez-Murcia AJ,
( 2008 ) Analysis of 16S rRNA gene mutations in a subset of Aeromonas strains and their impact in species delineation. PMID : 18843597 : Abstract >>
Characterization of 999 Aeromonas strains using a published 16S rDNA RFLP identification method showed that 8.1% of the strains produced unexpected (hereafter called "atypical") restriction patterns, making their identification uncertain. Atypical patterns were due to the presence of nucleotide polymorphisms among the rrn operons of the 16S rRNA gene (so-called microheterogeneities). Double sequencing signals at certain positions revealed the nucleotide composition was responsible for the microheterogeneities. Although the number of microheterogeneities was relatively low (0.06-0.66%), trees inferred from the 16S rRNA gene led either to a misidentification or to an inconclusive result for the majority of these strains. Strains with atypical patterns were, however, correctly identified using the rpoD gene sequences, as belonging to Aeromonas caviae, A. veronii, and A. media. All of them, but particularly the two former species, are associated with human disease. Microheterogeneities in 16S rRNA gene sequence were significantly (P 0.01) more prevalent in clinical than in environmental strains. This work also analyzed the effects of these microheterogeneities on the taxonomic position of the investigated strains. The results suggest the need for recording microheterogeneities in the 16S rRNA gene.
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15. |
Wan X,
Chai B,
Liao Y,
Su Y,
Ye T,
Shen P,
Chen X,
( 2009 ) Molecular and biochemical characterization of a distinct tyrosinase involved in melanin production from Aeromonas media. PMID : 18931836 : DOI : 10.1007/s00253-008-1742-5 Abstract >>
A new tyrosinase was isolated from Aeromonas media strain WS and purified to homogeneity. The purified tyrosinase, termed TyrA, had a molecular mass of 58 kDa and an isoelectric point of 4.90. It exhibited optimal monophenol and diphenol oxidase activities under basic conditions (pH>8.0). TyrA had a relatively higher affinity to diphenol substrate L-dihydroxyphenylalanine (L-dopa) than many other tyrosinases. EDTA or glutathione notably inhibited the enzymatic activities of TyrA, whereas Triton X-100 and SDS activated them. The full-length TyrA gene was cloned, and it encodes a 518 amino acid protein with little similarities to other reported tyrosinases. However, the purified recombinant TyrA expressed in Escherichia coli demonstrated tyrosinase activity. These results suggest that TyrA is the first reported distinct tyrosinase involved in melanin production in the genus Aeromonas.
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16. |
Sepe A,
Barbieri P,
Peduzzi R,
Demarta A,
( 2008 ) Evaluation of recA sequencing for the classification of Aeromonas strains at the genotype level. PMID : 18346137 : DOI : 10.1111/j.1472-765X.2008.02339.x Abstract >>
To evaluate the usefulness of partial recA sequences for the identification of Aeromonas strains at the genotype level. A partial recA sequence was obtained from 21 type or reference strains and 33 Aeromonas isolates, collected in the South of Switzerland from human, animal and aquatic environments. The 272 bp long recA fragments showed a mean interspecies divergence of 7.8% and allowed the classification of strains at genotype level. However, some discrepancies could be observed with other gene sequence based analyses in the classification of some strains. The 272 bp long recA fragment is a good molecular marker to infer taxonomy of members of the genus Aeromonas, even if the primers we chose for the amplification did not allow its direct sequencing. In the genus Aeromonas, nucleotide sequences of some protein-encoding genes have already been evaluated as molecular markers to be used in taxonomical and epidemiological researches. This study suggests the usefulness of a recA fragment as a further sequence to investigate for these purposes.
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17. |
Cattoir V,
Poirel L,
Aubert C,
Soussy CJ,
Nordmann P,
( 2008 ) Unexpected occurrence of plasmid-mediated quinolone resistance determinants in environmental Aeromonas spp. PMID : 18258115 : DOI : 10.3201/eid1402.070677 PMC : PMC2600179 Abstract >>
We searched for plasmid-mediated quinolone resistance determinants of the Qnr type in several water samples collected at diverse locations from the Seine River (Paris, France). The qnrS2 genes were identified from Aeromonas punctata subsp. punctata and A. media. The qnrS2 gene was located on IncU-type plasmids in both isolates, which resulted in increased MIC values of quinolones and fluoroquinolones, once they were transferred into Escherichia coli. The qnrS2 gene identified in A. punctata was part of novel genetic structure corresponding to a mobile insertion cassette element. This identification of plasmid-mediated qnr genes outside Enterobacteriaceae underlines a possible diffusion of those resistance determinants within gram-negative rods.
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18. |
Moura A,
Henriques I,
Ribeiro R,
Correia A,
( 2007 ) Prevalence and characterization of integrons from bacteria isolated from a slaughterhouse wastewater treatment plant. PMID : 17913715 : DOI : 10.1093/jac/dkm340 Abstract >>
To investigate the presence and distribution of integron-carrying bacteria from a slaughterhouse wastewater treatment plant (WWTP). Enterobacteriaceae and aeromonads were isolated at different stages of the wastewater treatment process and screened for the presence of integrase genes by dot-blot hybridization. Integrase-positive strains were characterized in terms of phylogenetic affiliation, genetic content of integrons and antimicrobial resistance profiles. Plasmid location of some integrons was established by Southern-blot hybridization. Strains containing integron-carrying plasmids were selected for mating experiments. Integrase genes were present in all samples, including the final effluent. The global prevalence was determined to be 35%, higher than in other aquatic environments. Forty-two integrase-positive isolates were further characterized. Nine distinct cassette arrays were found, containing genes encoding resistance to beta-lactams (bla(OXA-30)), aminoglycosides (aadA1, aadA2, aadA13, aadB), streptothricin (sat1, sat2), trimethoprim (dfrA1, dfrA12), a putative esterase (estX) and a protein with unknown function (orfF). Gene cassette arrays aadA1, dfrAI-aadA1 and estX-sat2-aadA1 were common to aeromonads and Enterobacteriaceae. The class 2 integron containing an estX-sat2-aadA1 cassette array was detected for the first time in Aeromonas sp. Nearly 12% (5 out of 43) of intI genes were located in plasmids. intI genes from isolates MM.1.3 and MM.1.5 were successfully conjugated into Escherichia coli at frequencies of 3.79 x 10(-5) and 5.46 x 10(-5) per recipient cell, respectively. Our data support the hypothesis that WWTPs constitute a potential hot spot for horizontal gene transfer and for selection of antimicrobial resistance genes among aquatic bacteria. Moreover, water discharges represent a possible risk for dissemination of undesirable genetic traits.
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19. |
Khor WC,
Puah SM,
Tan JA,
Puthucheary SD,
Chua KH,
( 2015 ) Phenotypic and Genetic Diversity of Aeromonas Species Isolated from Fresh Water Lakes in Malaysia. PMID : 26710336 : DOI : 10.1371/journal.pone.0145933 PMC : PMC4692508 Abstract >>
Gram-negative bacilli of the genus Aeromonas are primarily inhabitants of the aquatic environment. Humans acquire this organism from a wide range of food and water sources as well as during aquatic recreational activities. In the present study, the diversity and distribution of Aeromonas species from freshwater lakes in Malaysia was investigated using glycerophospholipid-cholesterol acyltransferase (GCAT) and RNA polymerase sigma-factor (rpoD) genes for speciation. A total of 122 possible Aeromonas strains were isolated and confirmed to genus level using the API20E system. The clonality of the isolates was investigated using ERIC-PCR and 20 duplicate isolates were excluded from the study. The specific GCAT-PCR identified all isolates as belonging to the genus Aeromonas, in agreement with the biochemical identification. A phylogenetic tree was constructed using the rpoD gene sequence and all 102 isolates were identified as: A. veronii 43%, A. jandaei 37%, A. hydrophila 6%, A. caviae 4%, A. salmonicida 2%, A. media 2%, A. allosaccharophila 1%, A. dhakensis 1% and Aeromonas spp. 4%. Twelve virulence genes were present in the following proportions--exu 96%, ser 93%, aer 87%, fla 83%, enolase 70%, ela 62%, act 54%, aexT 33%, lip 16%, dam 16%, alt 8% and ast 4%, and at least 2 of these genes were present in all 102 strains. The ascV, aexU and hlyA genes were not detected among the isolates. A. hydrophila was the main species containing virulence genes alt and ast either present alone or in combination. It is possible that different mechanisms may be used by each genospecies to demonstrate virulence. In summary, with the use of GCAT and rpoD genes, unambiguous identification of Aeromonas species is possible and provides valuable data on the phylogenetic diversity of the organism.
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20. |
Mosser T,
Talagrand-Reboul E,
Colston SM,
Graf J,
Figueras MJ,
Jumas-Bilak E,
Lamy B,
( 2015 ) Exposure to pairs of Aeromonas strains enhances virulence in the Caenorhabditis elegans infection model. PMID : 26583012 : DOI : 10.3389/fmicb.2015.01218 PMC : PMC4631986 Abstract >>
Aeromonad virulence remains poorly understood, and is difficult to predict from strain characteristics. In addition, infections are often polymicrobial (i.e., are mixed infections), and 5-10% of such infections include two distinct aeromonads, which has an unknown impact on virulence. In this work, we studied the virulence of aeromonads recovered from human mixed infections. We tested them individually and in association with other strains with the aim of improving our understanding of aeromonosis. Twelve strains that were recovered in pairs from six mixed infections were tested in a virulence model of the worm Caenorhabditis elegans. Nine isolates were weak worm killers (median time to death, TD50, ?7 days) when administered alone. Two pairs showed enhanced virulence, as indicated by a significantly shortened TD50 after co-infection vs. infection with a single strain. Enhanced virulence was also observed for five of the 14 additional experimental pairs, and each of these pairs included one strain from a natural synergistic pair. These experiments indicated that synergistic effects were frequent and were limited to pairs that were composed of strains belonging to different species. The genome content of virulence-associated genes failed to explain virulence synergy, although some virulence-associated genes that were present in some strains were absent from their companion strain (e.g., T3SS). The synergy observed in virulence when two Aeromonas isolates were co-infected stresses the idea that consideration should be given to the fact that infection does not depend only on single strain virulence but is instead the result of a more complex interaction between the microbes involved, the host and the environment. These results are of interest for other diseases in which mixed infections are likely and in particular for water-borne diseases (e.g., legionellosis, vibriosis), in which pathogens may display enhanced virulence in the presence of the right partner. This study contributes to the current shift in infectiology paradigms from a premise that assumes a monomicrobial origin for infection to one more in line with the current pathobiome era.
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21. |
Persson S,
Al-Shuweli S,
Yapici S,
Jensen JN,
Olsen KE,
( 2015 ) Identification of clinical aeromonas species by rpoB and gyrB sequencing and development of a multiplex PCR method for detection of Aeromonas hydrophila, A. caviae, A. veronii, and A. media. PMID : 25411168 : DOI : 10.1128/JCM.01963-14 PMC : PMC4298543 Abstract >>
Conventional identification of Aeromonas species based on biochemical methods is challenged by the heterogeneous nature of the species. Here, we present a new multiplex PCR method directed toward the gyrB and rpoB genes that identifies four Aeromonas species, A. hydrophila, A. media, A. veronii, and A. caviae, and we describe the application of this method on a Danish strain collection.
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22. |
Vega-Sánchez V,
Latif-Eugenín F,
Soriano-Vargas E,
Beaz-Hidalgo R,
Figueras MJ,
Aguilera-Arreola MG,
Castro-Escarpulli G,
( 2014 ) Re-identification of Aeromonas isolates from rainbow trout and incidence of class 1 integron and �]-lactamase genes. PMID : 25008317 : DOI : 10.1016/j.vetmic.2014.06.012 Abstract >>
Forty-eight Aeromonas isolates from rainbow trout previously identified by the 16S rDNA-RFLP technique were re-identified using 2 housekeeping genes (gyrB and rpoD). After sequencing the prevalences of the species were A. veronii (29.2%), A. bestiarum (20.8%), A. hydrophila (16.7%), A. sobria (10.4%), A. media (8.3%), A. popoffii (6.2%), A. allosaccharophila (2.1%), A. caviae (2.1%), A. salmonicida (2.1%) and one isolate (2.1%) belongs to a candidate new species "Aeromonas lusitana". Coincident identification results to the 16S rDNA-RFLP technique were only obtained for 68.8% of the isolates. PCR amplification of the enterobacterial repetitive intergenic consensus (ERIC-PCR) indicated that the 48 isolates belonged to 33 different ERIC genotypes. Several genotypes were isolated from different farms and organs in the same fish, indicating a systemic dissemination of the bacteria. The presence of genes (blaIMP, blaCphA/IMIS, blaTEM, blaSHV and intI1) that encode extended-spectrum beta-lactamases (ESBLs), metallo-beta-lactamases (MBLs) and class 1 integrons were studied by PCR. Only 39.6% (19/48) of the strains showed the presence of one or more resistance genes. The gene blaCphA/IMIS was detected in 29.2% of the isolates, followed by the intI1 (6.2%) and blaSHV (4.2%) genes. The variable region of class 1 integrons of the 3 positive isolates was sequenced revealing the presence of the gene cassette aadA1 (aminoglycoside transferase) that plays a role in streptomycin/spectinomycin resistance.
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23. |
Martino ME,
Fasolato L,
Montemurro F,
Novelli E,
Cardazzo B,
( 2014 ) Aeromonas spp.: ubiquitous or specialized bugs? PMID : 23919504 : DOI : 10.1111/1462-2920.12215 Abstract >>
The genus Aeromonas comprises ubiquitous bacteria that are known to play several roles in the environment. These bacteria were first described as fish pathogens, but their presence was documented in other reservoirs, such as animals and humans. Today, these bacteria are described as emerging pathogens, but their effective role in human pathogenicity is still controversial. In addition, their taxonomy is heavily debated, as species distinction is often difficult to achieve. To study the interspecies relationships and to investigate their connection with the environment, a multilocus sequence typing scheme previously developed for Aeromonas spp. was applied to 258 strains, and the genetic data were analysed by population software. Sampling was a fundamental step, including several of the main sources of Aeromonas: fish, food products and human cases of disease. The objective was to characterize the isolates and to find potential associations among them according to the following: species, sharing of virulence factors, source and adaptation to a specific habitat. The strains were characterized and demonstrated exceptionally high nucleotide variability in the Aeromonas genus. Among the sampled sources, different species distributions were found, highlighting the occurrence of adaptation processes towards specific habitats.
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24. |
Roger F,
Marchandin H,
Jumas-Bilak E,
Kodjo A,
N/A N/A,
Lamy B,
( 2012 ) Multilocus genetics to reconstruct aeromonad evolution. PMID : 22545815 : DOI : 10.1186/1471-2180-12-62 PMC : PMC3487998 Abstract >>
Aeromonas spp. are versatile bacteria that exhibit a wide variety of lifestyles. In an attempt to improve the understanding of human aeromonosis, we investigated whether clinical isolates displayed specific characteristics in terms of genetic diversity, population structure and mode of evolution among Aeromonas spp. A collection of 195 Aeromonas isolates from human, animal and environmental sources was therefore genotyped using multilocus sequence analysis (MLSA) based on the dnaK, gltA, gyrB, radA, rpoB, tsf and zipA genes. The MLSA showed a high level of genetic diversity among the population, and multilocus-based phylogenetic analysis (MLPA) revealed 3 major clades: the A. veronii, A. hydrophila and A. caviae clades, among the eleven clades detected. Lower genetic diversity was observed within the A. caviae clade as well as among clinical isolates compared to environmental isolates. Clonal complexes, each of which included a limited number of strains, mainly corresponded to host-associated subsclusters of strains, i.e., a fish-associated subset within A. salmonicida and 11 human-associated subsets, 9 of which included only disease-associated strains. The population structure was shown to be clonal, with modes of evolution that involved mutations in general and recombination events locally. Recombination was detected in 5 genes in the MLSA scheme and concerned approximately 50% of the STs. Therefore, these recombination events could explain the observed phylogenetic incongruities and low robustness. However, the MLPA globally confirmed the current systematics of the genus Aeromonas. Evolution in the genus Aeromonas has resulted in exceptionally high genetic diversity. Emerging from this diversity, subsets of strains appeared to be host adapted and/or disease specialized" while the A. caviae clade displayed an atypical tempo of evolution among aeromonads. Considering that A. salmonicida has been described as a genetically uniform pathogen that has adapted to fish through evolution from a variable ancestral population, we hypothesize that the population structure of aeromonads described herein suggested an ongoing process of adaptation to specialized niches associated with different degrees of advancement according to clades and clusters."
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25. |
Pan ZH,
Lu CP,
( 2012 ) Identity and virulence properties of Aeromonas isolates from diseased fish, healthy controls and water environment in China. PMID : 22725694 : DOI : 10.1111/j.1472-765X.2012.03281.x Abstract >>
To investigate the species distribution in Aeromonas isolates from diseased fish, healthy controls and water environment in China; to evaluate the frequency of the aerolysin (aer), cytotonic enterotoxin (alt), cytotoxic enterotoxin (act), temperature-sensitive protease (eprCAI) and serine protease (ahp) genes in Aeromonas isolates; and to determine the potential pathogenicity of these isolates. Two hundred and two Aeromonas isolates from diseased fish (n = 42), healthy fish (n = 120) and water environment (n = 40) in China were identified to species levels based on sequencing of the housekeeping gene gyrB, while the distribution of five virulence factors, including aer, alt, act, eprCAI and ahp, was investigated by PCR. Aeromonas veronii (25/42; 60%) and Aeromonas hydrophila (14/42; 33%) were the species most commonly isolated from diseased fish, while Aer. veronii was the most common species in healthy fish (90/120; 75%) and water samples (25/40; 62�P5%). All the five virulence genes were present in 9% (19/202), among which 10 strains were from diseased fish and nine were identified as Aer. hydrophila. For the strains carrying five virulence genes, the average 50% lethal doses (LD(50s)) of strains from diseased fish were lower when compared with the strains from healthy fish and water environment. Aeromonas veronii is the most common species, but no significant difference exists in the isolates obtained from diseased fish and from healthy fish. However, Aer. hydrophila isolates were significantly more frequent from diseased fish than from healthy fish. aer+alt+act+eprCAI+ ahp+ was more frequent virulence genotype in Aeromonas isolates from diseased fish than from healthy fish and water environment, and the aer+alt+act+eprCAI+ahp+ isolates were more virulent to zebrafish comparing to the other genetic profiles. Aeromonas species in aquatic environments are various and have considerable virulence potential, and therefore, there is a need for more careful and intensive epidemiology studies.
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26. |
Martino ME,
Fasolato L,
Montemurro F,
Rosteghin M,
Manfrin A,
Patarnello T,
Novelli E,
Cardazzo B,
( 2011 ) Determination of microbial diversity of Aeromonas strains on the basis of multilocus sequence typing, phenotype, and presence of putative virulence genes. PMID : 21642403 : DOI : 10.1128/AEM.00708-11 PMC : PMC3147379 Abstract >>
The genus Aeromonas has been described as comprising several species associated with the aquatic environment, which represents their principal reservoir. Aeromonas spp. are commonly isolated from diseased and healthy fish, but the involvement of such bacteria in human infection and gastroenteritis has frequently been reported. The primary challenge in establishing an unequivocal link between the Aeromonas genus and pathogenesis in humans is the extremely complicated taxonomy. With the aim of clarifying taxonomic relationships among the strains and phenotypes, a multilocus sequencing approach was developed and applied to characterize 23 type and reference strains of Aeromonas spp. and a collection of 77 field strains isolated from fish, crustaceans, and mollusks. All strains were also screened for putative determinants of virulence by PCR (ast, ahh1, act, asa1, eno, ascV, and aexT) and the production of acylated homoserine lactones (AHLs). In addition, the phenotypic fingerprinting obtained from 29 biochemical tests was submitted to the nonparametric combination (NPC) test methodology to define the statistical differences among the identified genetic clusters. Multilocus sequence typing (MLST) achieved precise strain genotyping, and the phylogenetic analysis of concatenated sequences delineated the relationship among the taxa belonging to the genus Aeromonas, providing a powerful tool for outbreak traceability, host range diffusion, and ecological studies. The NPC test showed the feasibility of phenotypic differentiation among the majority of the MLST clusters by using a selection of tests or the entire biochemical fingerprinting. A Web-based MLST sequence database (http://pubmlst.org/aeromonas) specific for the Aeromonas genus was developed and implemented with all the results.
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27. |
Moura A,
Pereira C,
Henriques I,
Correia A,
( 2012 ) Novel gene cassettes and integrons in antibiotic-resistant bacteria isolated from urban wastewaters. PMID : 22127350 : DOI : 10.1016/j.resmic.2011.10.010 Abstract >>
In this study, the occurrence and diversity of integrons were evaluated in 697 isolates belonging to Enterobacteriaceae and Aeromonas spp. isolated from urban wastewaters. Screening of integrons was performed by dot blot hybridization and intI-positive strains were further characterized. The global prevalence of integrons was 3.73%. Three new gene cassettes were identified: a novel aadA variant (aadA17), a gene putatively involved in cell signaling (dcyA) and an open reading frame of unknown function interrupted by a novel insertion sequence (orfER.17::ISAs12). In total, thirteen different gene cassette arrays were detected, 4 representing novel integrons: intI1-dcyA-tniC, intI1-orfER.1.7::ISAs12-aadA13-qacE�G1-sul1, intI1-aacA4-catB3-bla(OxA-10)-aadA1-qacE�G1-sul1 and intI1-catB8-aadA17-qacE�G1-sul1. Approximately 80% of strains were resistant to at least 3 antibiotics of different classes. The presence of novel integron structures in treated effluents suggests that domestic wastewaters may favor the formation of novel combinations of gene cassettes. Moreover, the high prevalence of multiresistant strains highlights the urgent need to employ effective means of effluent disinfection to avoid dissemination of antibiotic-resistant bacteria.
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28. |
Amos GCA,
Ploumakis S,
Zhang L,
Hawkey PM,
Gaze WH,
Wellington EMH,
( 2018 ) The widespread dissemination of integrons throughout bacterial communities in a riverine system. PMID : 29374269 : DOI : 10.1038/s41396-017-0030-8 PMC : PMC5864220 Abstract >>
Anthropogenic inputs increase levels of antimicrobial resistance (AMR) in the environment, however, it is unknown how these inputs create this observed increase, and if anthropogenic sources impact AMR in environmental bacteria. The aim of this study was to characterise the role of waste water treatment plants (WWTPs) in the dissemination of class 1 integrons (CL1s) in the riverine environment. Using sample sites from upstream and downstream of a WWTP, we demonstrate through isolation and culture-independent analysis that WWTP effluent significantly increases both CL1 abundance and antibiotic resistance in the riverine environment. Characterisation of CL1-bearing isolates revealed that CL1s were distributed across a diverse range of bacteria, with identical complex genetic resistance determinants isolated from both human-associated and common environmental bacteria across connected sites. Over half of sequenced CL1s lacked the 3'-conserved sequence ('atypical' CL1s); surprisingly, bacteria carrying atypical CL1s were on average resistant to more antibiotics than bacteria carrying 3'-CS CL1s. Quaternary ammonium compound (QAC) resistance genes were observed across 75% of sequenced CL1 gene cassette arrays. Chemical data analysis indicated high levels of boron (a detergent marker) downstream of the WWTP. Subsequent phenotypic screening of CL1-bearing isolates demonstrated that ~90% were resistant to QAC detergents, with in vitro experiments demonstrating that QACs could solely select for the transfer of clinical antibiotic resistance genes to a naive Escherichia coli recipient. In conclusion, this study highlights the significant impact of WWTPs on environmental AMR, and demonstrates the widespread carriage of clinically important resistance determinants by environmentally associated bacteria.
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29. |
Hoel S,
Vadstein O,
Jakobsen AN,
( 2017 ) Species Distribution and Prevalence of Putative Virulence Factors in Mesophilic Aeromonas spp. Isolated from Fresh Retail Sushi. PMID : 28596762 : DOI : 10.3389/fmicb.2017.00931 PMC : PMC5442234 Abstract >>
Aeromonas spp. are ubiquitous bacteria that have received increasing attention as human pathogens because of their widespread occurrence in food, especially seafood and vegetables. The aim of this work was to assess the species identity and phylogenetic relationship of 118 Aeromonas strains isolated from fresh retail sushi from three producers, and to characterize the isolates with respect to genetic and phenotypic virulence factors. We also evaluate the potential hazard associated with their presence in ready-to-eat seafood not subjected to heat treatment. Mesophilic Aeromonas salmonicida was most prevalent (74%), followed by A. bestiarum (9%), A. dhakensis (5%), A. caviae (5%), A. media (4%), A. hydrophila (2%), and A. piscicola (1%). All isolates were considered potentially pathogenic due to the high prevalence of genes encoding hemolysin (hlyA) (99%), aerolysin (aerA) (98%), cytotoxic enterotoxin (act) (86%), heat-labile cytotonic enterotoxin (alt) (99%), and heat-stable cytotonic enterotoxin (ast) (31%). The shiga-like toxins 1 and 2 (stx-1 and stx-2) were not detected. Moreover, there was heterogeneity in toxin gene distribution among the isolates, and the combination of act/alt/hlyA/aerA was most commonly detected (63%). �]-hemolysis was species-dependent and observed in 91% of the isolates. All A. media and A. caviae strains were non-hemolytic. For isolates belonging to this group, lack of hemolysis was possibly related to the absence of the act gene. Swimming motility, linked to adhesion and host invasion, occurred in 65% of the isolates. Partial sequencing of the gyrB gene demonstrated its suitability as a genetic marker for Aeromonas species identification and for assessment of the phylogenetic relationship between the isolates. The gyrB sequence divergence within a given species ranged from 1.3 to 2.9%. A. bestiarum, A. salmonicida, and A. piscicola were the most closely related species; their sequences differed by 2.7-3.4%. The average gyrB sequence similarity between all species was 93%, demonstrating its acceptable taxonomic resolution. The presence of multiple species of potential pathogenic Aeromonas in fresh retail sushi raises new food safety issues related to the increased consumption of ready-to-eat food composed of raw ingredients.
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30. |
( 2012 ) Phylogenetic diversity, antibiotic resistance and virulence traits of Aeromonas spp. from untreated waters for human consumption. PMID : 23107502 : DOI : 10.1016/j.ijfoodmicro.2012.09.008 Abstract >>
It is well known that water constitutes an important contamination route for microorganisms. This is especially true for Aeromonas which are widespread in untreated and treated waters. In this study, Portuguese untreated waters not regularly monitored were screened for the presence and diversity of aeromonads. A total of 206 isolates were discriminated by RAPD-PCR and 80 distinct strains were identified by gyrB based phylogenetic analysis. The most frequently detected species were Aeromonas hydrophila, Aeromonas bestiarum and Aeromonas media. The antibiotic susceptibility profile of these strains was determined and showed a typical profile of the genus. Nonetheless, the percentage of resistant strains to tetracycline, chloramphenicol and/or trimethoprim/sulfamethoxazole was lower than that reported for clinical isolates and isolates recovered from aquacultures and other environments historically subjected to antibiotic contamination. This suggests that the existence of such pressures in those environments selects for resistant Aeromonas. A similar trend for integron presence was found. Genes coding for CphA and TEM, and tet(A), (E), (C) or (D) genes were found in 28%, 1%, and 10% of the strains, respectively. 10% of the strains contained an integron. Variable regions of seven class 1 integrons and one class 2 integron were characterised. Furthermore, strains displayed virulence related phenotypes such as extracellular lipolytic and proteolytic activities as well as aerolysin related genes (43% of strains). The ascV and aexT genes were found in 16% and 3% of strains respectively and, in some cases, concomitantly in the same specimen. This study shows that diverse Aeromonas spp. presenting distinct antibiotic resistance features and putative virulence traits are frequently present in waters for human and animal consumption in Portugal. Genes associated to antibiotic resistance and microbial virulence previously identified in organisms with human health significance were detected in these aeromonads, suggesting that these waters may act as a pivotal route for infections.
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31. |
( 2012 ) Changes in the gut microbiome of the sea lamprey during metamorphosis. PMID : 22923392 : DOI : 10.1128/AEM.01640-12 PMC : PMC3485723 Abstract >>
Vertebrate metamorphosis is often marked by dramatic morphological and physiological changes of the alimentary tract, along with major shifts in diet following development from larva to adult. Little is known about how these developmental changes impact the gut microbiome of the host organism. The metamorphosis of the sea lamprey (Petromyzon marinus) from a sedentary filter-feeding larva to a free-swimming sanguivorous parasite is characterized by major physiological and morphological changes to all organ systems. The transformation of the alimentary canal includes closure of the larval esophagus and the physical isolation of the pharynx from the remainder of the gut, which results in a nonfeeding period that can last up to 8 months. To determine how the gut microbiome is affected by metamorphosis, the microbial communities of feeding and nonfeeding larval and parasitic sea lamprey were surveyed using both culture-dependent and -independent methods. Our results show that the gut of the filter-feeding larva contains a greater diversity of bacteria than that of the blood-feeding parasite, with the parasite gut being dominated by Aeromonas and, to a lesser extent, Citrobacter and Shewanella. Phylogenetic analysis of the culturable Aeromonas from both the larval and parasitic gut revealed that at least five distinct species were represented. Phenotypic characterization of these isolates revealed that over half were capable of sheep red blood cell hemolysis, but all were capable of trout red blood cell hemolysis. This suggests that the enrichment of Aeromonas that accompanies metamorphosis is likely related to the sanguivorous lifestyle of the parasitic sea lamprey.
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