Fig 1.
Nezara viridula hosts survey around the year cycle.
Secondary hosts (SH) from late September to late January; soybean (primary host) from February to late May and under the bark of Eucalyptus trees (diapause) from June to late September.
Table 1.
Microscopic and molecular identification of 21 isolates from the midgut of field collected N. viridula adults.
Fig 2.
Presence/absence of gut microbiota in N. viridula adults associated with the insect host.
Presence of non-transient (black) and transient (light grey) microbiota or absence of bacteria (Not infected; dark grey) in N. viridula adult’s V1-V3 midgut ventricles; and its distribution related to insect host: secondary hosts (SH), soybean and under the bark of Eucalyptus trees. Numbers correspond to insect gut dissected.
Fig 3.
Gut bacterial communities associated with the insect host.
Bacteria species inhabiting Nezara viridula infected V1-V3 midgut ventricles and its distribution among insect hosts (a) and relative abundance (CFU/mg gut x n° insects positive -1) of bacterial groups of infected N. viridula V1-V3 midgut ventricles and its distribution among hosts (b). *Enterobacteriaceae groups Yokenella, Cedecea and Pantoea species. Numbers correspond to insect gut dissected.
Fig 4.
Phylogenetic placements of Yokenella strains (bold letters).
We constructed a tree that included our isolates, those described as K. pneumoniae by Hirose et al. (2006), the species reference strains K. pneumoniae ATCC 13885, K. michiganensis ATCC BAA-2403, K. oxytoca ATCC 13182, Y. regensburgei ATCC 49455 and one additional strain of Y. regensburgei. Gene bank accession numbers and insect host of bacteria isolated in this study appear in parentheses Escherichia coli ATCC 11775 was included as an outgroup to root the tree. K. michiganensis and K. oxytoca were included because a blast search in the RefSeq database indicates that both these species and Y. regensburgei lie at closer genetic distance to the Hirose´s strains than K. pneumoniae. Only the bootstrap values greater than 60% are shown. Scale bar states for the phylogenetic distance with a common ancestor.
Fig 5.
Phylogenetic positioning of Pantoea NvP01 (bold letters).
We constructed a tree that included our isolate and type strains of 22 species of Pantoea and a subspecie of Pantoea stewartii. Gene bank accession numbers and insect host of bacteria isolated in this study appear in parentheses. The sequence of the type strain Klebsiella pneumoniae ATCC 13883 was included as an outgroup to root the tree. Only the bootstrap values greater than 65% are shown. Scale bar states for the phylogenetic distance with a common ancestor.
Fig 6.
Phylogenetic positioning of Enterococcus isolated strains (bold letters).
We constructed a tree that included our isolates, those described as Enterococcus faecalis by Hirose et al. (2006), and type strain Enterococcus faecalis JCM 5803. Gene bank accession numbers and insect host of bacteria isolated in this study appear in parentheses. The sequence of the type strain Enterococcus moraviensis ATCC BAA-383 was included as an outgroup to root the tree. Only the bootstrap values greater than 65% are shown. Scale bar states for the phylogenetic distance with a common ancestor.
Table 2.
Enzymatic activities of isolated strains with significance in soybean digestion.
Fig 7.
Inhibition capacity of fermented soybean cv. Williams whole meal extracts against cysteine protease papain.
Whole meal was inoculated with non-transient microbiota (NTM) isolated bacteria and fermented during 24h at 37°C. Yokenella sp. NvH01, NvO01, NvR01, NvU01, NvU02 and NvW01; Pantoea sp. NvP01; Enterococcus faecalis NvH02 and Enterococcus p: NvM04. Control extracts correspond to a pasteurized non-inoculated and incubated soybean whole meal suspension.