Effects of temperature onParamoeba peruransgrowth in culture and the associated microbial community

Abstract

Population growth, in vitro, of three Paramoeba perurans cultures, one polyclonal (G) and two clonal (B8, CE6, derived from G), previously shown to differ in virulence (B8 > G > CE6), was compared at 10 and 15 °C. B8 showed a significantly higher increase in attached and in suspended amoebae over time at 15 and 10 °C, respectively. CE6 and G also had significantly higher numbers of suspended amoebae at 10 °C compared with 15 °C at experiment termination. However, in contrast to B8, numbers of attached amoebae were significantly higher at 10 °C in CE6 but showed a similar trend in G at the end of the experiment. Numbers of both suspended and attached amoebae were lower in B8 compared with CE6 and G. Significant differences in bacterial community composition and/or relative abundances were found, between cultures, between temperatures and between the same culture with and without amoebae, based on 16S rRNA Illumina MiSeq sequencing. Bacterial diversity was lower in B8 and CE6 compared with G, possibly reflecting selection during clonal isolation. The results indicate that polyclonal P. perurans populations may contain amoebae displaying different growth dynamics. Further studies are required to determine if these differences are linked to differences seen in the bacterial communities.

Effects of temperature on amoebic gill disease development: Does it play a role?

“…Similar letters indicate that different treatments were not statistically different (p > 0.05, n = 40) a significant difference between infected fish held at 10 and 15°C during the first, second and third samplings after P. perurans exposure with an earlier detection and higher numbers (or possibly higher expression activity) of P. perurans associated with the higher temperature (15°C). A recent study using the same P. perurans clone as in this study (B8) showed in vitro a significantly higher increase in attached amoeba over time at 15°C than at 10°C (while amoebae in suspension increased to a greater extent at 10°C), and this phenomenon perhaps contributes to the findings here (Benedicenti et al, 2019).…”

“…Immune-and stress-related gene expression analysis showed modulation in gills rather than in head kidney, mainly during the first sampling point after challenge, with different expression levels between the two organs revealing an organ-specific response. Therefore, higher temperature (at 15°C) while linked mainly to earlier and stronger P. perurans infection through supporting greater proliferation of P. perurans on gills, at least for the amoeba clone used here (Benedicenti et al, 2019), may also act as a potential stressor in terms of changes in hormone levels in the plasma during early stages of pathology.…”

“…Negative and control groups are described in Table 1. The amoeba cultures were acclimated at the two different temperatures (10 and 15°C) for 3 months before starting the experiment, and four passages of the cultures were performed over this time to maintain their survival until the start of the experiment as described by Benedicenti, Secombes, and Collins (2019). Amoebae were cultured as described previously (Benedicenti, Collins, Wang, McCarthy, & Secombes, 2015), and aliquots of amoebae were numerically equal and randomly distributed to experimental tanks.…”

Comparative proteomic profiling of newly acquired, virulent and attenuated Neoparamoeba perurans proteins associated with amoebic gill disease

“…Due to the abundance of literature suggesting N. perurans microbiome may in uence AGD progression 26,27,29,30 , 16S rRNA sequencing was employed to clarify the role of the microbiome in the virulent and attenuated isolates. Fig.…”

“…Several genera coupled to Pseudomonas spp, Marinomonas sp and Flavobacterium sp were identi ed 29 . Polygonal and clonal isolates of N. perurans shared several genera of both gram positive and gram negative bacteria with signi cant differences in relative abundances between the three isolates 30 . Despite the speculation surrounding the role of N. perurans microbiome in AGD, no study exists that directly compares the microbiome of a long-term cultured N. perurans isolate to a virulent isolate.…”