MOESM1 of RNA-Seq based transcriptome of whole blood from immunocompetent pigs (Sus scrofa) experimentally infected with Mycoplasma suis strain Illinois

Additional file 1. Results of Mycoplasma suis microbead immunoassay (MIA). Median Fluorescence Intensity (MFI) of Mycoplasma suis-infected and sham-inoculated pigs as detected in serum samples collected along the experiment. Serum antibodies were measured using recombinant GrpE (rGrpE) protein in a MIA [30]. Sham-inoculated group (control group): pig #1, pig #2 and pig #3. M. suis-inoculated group (infected group): pig #5, pig #6, pig #7. Day 0 corresponds to inoculation day. MFI cut-off was 74.07. It is important to note that a high cut-off may have been encountered due to the low number of animals to determine a cut-off, which increases the standard deviation. MFI values in bold correspond to positive results based on cut-off values for each group. ND: not done

MOESM2 of RNA-Seq based transcriptome of whole blood from immunocompetent pigs (Sus scrofa) experimentally infected with Mycoplasma suis strain Illinois

Additional file 2. Results of RNA-Seq analyses. List of differentially expressed (DE) genes identified in whole blood of Mycoplasma suis-infected pigs (when compared to sham-infected group) separated by all three methods of RNA-Seq analyses (DESeq2, edgeR, Cufflinks). First tab describes all DE genes (p-value ≤ 0.1) that were detected in all three methods, in two methods, followed by each individual method. The following tabs report the statistics of each evaluated gene for each method. Highlighted in grey are statistically significant DE genes (dark grey: p-value ≤ 0.05; light grey p-value ≤ 0.1)

MOESM3 of RNA-Seq based transcriptome of whole blood from immunocompetent pigs (Sus scrofa) experimentally infected with Mycoplasma suis strain Illinois

Additional file 3. Relative expression (RE) of differentially expressed (DE) in pigs infected with Mycoplasma suis validated by qRT-PCR. Relative expression profile (GAPDH as reference control gene) of DE genes identified in the M. suis-infected pigs compared to the control group (non-infected). Validated genes are: ISG15 ubiquitin-like modifier (ISG15), interleukin 22 receptor, alpha 2 (IL22RA2), CD274 molecule (CD274), BCL2-like 14 (apoptosis facilitator) (BCL2), chemokine (C-C motif) receptor 5 (CCR5), CD180 molecule (CD180), allograft inflammatory factor 1 (AIF1), interleukin 15 (IL15), protein tyrosine phosphatase, receptor type, O (PTPRO), and Toll-like receptor 8 (TLR8). Black dots in each graphic represent the RE of a respective gene for each pig; blue vertical lines represent the standard deviation in each group. * p-value ≤ 0.1, and ** p-value ≤ 0.05

MOESM4 of RNA-Seq based transcriptome of whole blood from immunocompetent pigs (Sus scrofa) experimentally infected with Mycoplasma suis strain Illinois

Additional file 4. Gene ontology (GO) categories of all 55 differentially expressed (DE) genes of Mycoplasma suis infected pigs. First tab describes raw results of GO analysis, with all DE genes and their correspondent GO categories (and functions). Second tab shows the identified functions of the “biological process” category of all 55 DE genes and the number of reported genes for each function. Functional categories with the largest number of genes are highlighted in yellow. GO analysis was performed using Blast2Go software [39]

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<p>Mycoplasma bovis is a major bovine pathogen that causes considerable economic losses in the cattle industry worldwide. Moreover, M. bovis biofilm can persist in the environment and its host. To date, M. bovis biofilm antigens recognized by bovine convalescent sera and their comparison with planktonic cells have not yet been explored. This study utilized an immunoproteomic approach using two-dimensional electrophoresis, immunoblotting using convalescent bovine serum, and subsequent matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry (MALDI-TOF/TOF MS) to identify the immunoreactive proteins expressed in biofilm- and planktonic-grown M. bovis strain 08M. Results showed that M. bovis biofilms and planktonic cells demonstrate differential immunoreactivity to bovine convalescent serum for the first time. A total of 10 and 8 immunoreactive proteins were identified for biofilms and planktonic cells, respectively. To our knowledge, a total of 12 out of 15 had not been reported as immunoreactive proteins in M. bovis, and six were specific to M. bovis biofilms. Three proteins, namely, endoglucanase, thiol peroxidase, and one putative membrane protein, that is, mycoplasma immunogenic lipase A, were identified in planktonic cells and biofilms. Most of the identified proteins were cytoplasmic proteins that were mainly involved in transport and metabolism. Moreover, ATP binding, oxidoreductase activity, and GTP binding were their most representative molecular functions. DnaK and Tuf appeared to be the most interactive immunoreactive agent among the identified proteins. Furthermore, six proteins had potential as serodiagnostic antigens. These data will be helpful to improve our current understanding on the host response to M. bovis biofilms and planktonic cells, which may facilitate the development of novel molecular candidates of improved diagnostics and vaccines to prevent M. bovis infections.</p

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<p>Mycoplasma bovis is a major bovine pathogen that causes considerable economic losses in the cattle industry worldwide. Moreover, M. bovis biofilm can persist in the environment and its host. To date, M. bovis biofilm antigens recognized by bovine convalescent sera and their comparison with planktonic cells have not yet been explored. This study utilized an immunoproteomic approach using two-dimensional electrophoresis, immunoblotting using convalescent bovine serum, and subsequent matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry (MALDI-TOF/TOF MS) to identify the immunoreactive proteins expressed in biofilm- and planktonic-grown M. bovis strain 08M. Results showed that M. bovis biofilms and planktonic cells demonstrate differential immunoreactivity to bovine convalescent serum for the first time. A total of 10 and 8 immunoreactive proteins were identified for biofilms and planktonic cells, respectively. To our knowledge, a total of 12 out of 15 had not been reported as immunoreactive proteins in M. bovis, and six were specific to M. bovis biofilms. Three proteins, namely, endoglucanase, thiol peroxidase, and one putative membrane protein, that is, mycoplasma immunogenic lipase A, were identified in planktonic cells and biofilms. Most of the identified proteins were cytoplasmic proteins that were mainly involved in transport and metabolism. Moreover, ATP binding, oxidoreductase activity, and GTP binding were their most representative molecular functions. DnaK and Tuf appeared to be the most interactive immunoreactive agent among the identified proteins. Furthermore, six proteins had potential as serodiagnostic antigens. These data will be helpful to improve our current understanding on the host response to M. bovis biofilms and planktonic cells, which may facilitate the development of novel molecular candidates of improved diagnostics and vaccines to prevent M. bovis infections.</p

Image_2.TIF

<p>Mycoplasma bovis is a major bovine pathogen that causes considerable economic losses in the cattle industry worldwide. Moreover, M. bovis biofilm can persist in the environment and its host. To date, M. bovis biofilm antigens recognized by bovine convalescent sera and their comparison with planktonic cells have not yet been explored. This study utilized an immunoproteomic approach using two-dimensional electrophoresis, immunoblotting using convalescent bovine serum, and subsequent matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry (MALDI-TOF/TOF MS) to identify the immunoreactive proteins expressed in biofilm- and planktonic-grown M. bovis strain 08M. Results showed that M. bovis biofilms and planktonic cells demonstrate differential immunoreactivity to bovine convalescent serum for the first time. A total of 10 and 8 immunoreactive proteins were identified for biofilms and planktonic cells, respectively. To our knowledge, a total of 12 out of 15 had not been reported as immunoreactive proteins in M. bovis, and six were specific to M. bovis biofilms. Three proteins, namely, endoglucanase, thiol peroxidase, and one putative membrane protein, that is, mycoplasma immunogenic lipase A, were identified in planktonic cells and biofilms. Most of the identified proteins were cytoplasmic proteins that were mainly involved in transport and metabolism. Moreover, ATP binding, oxidoreductase activity, and GTP binding were their most representative molecular functions. DnaK and Tuf appeared to be the most interactive immunoreactive agent among the identified proteins. Furthermore, six proteins had potential as serodiagnostic antigens. These data will be helpful to improve our current understanding on the host response to M. bovis biofilms and planktonic cells, which may facilitate the development of novel molecular candidates of improved diagnostics and vaccines to prevent M. bovis infections.</p

Characterization of Chinese <i>Haemophilus parasuis</i> Isolates by Traditional Serotyping and Molecular Serotyping Methods

<div><p><i>Haemophilus parasuis</i> is classified mainly through serotyping, but traditional serotyping always yields non-typable (NT) strains and unreliable results via cross-reactions. Here, we surveyed the serotype prevalence of Chinese <i>H</i>. <i>parasuis</i> isolates using traditional serotyping (gel immuno-diffusion test, GID) and molecular serotyping (multiplex PCR, mPCR). We also investigated why discrepant results between these methods were obtained, and investigated mPCR failure through whole-genome sequencing. Of the 100 isolate tested, 73 (73%) and 93 (93%) were serotyped by the GID test and mPCR, respectively, with a concordance rate of 66% (66/100). Additionally, mPCR reduced the number of NT isolates from 27 (27%) for the GID testing, to seven (7%). Eleven isolates were sequenced, including nine serotype-discrepant isolates from mPCR and GID typing (excluding strains that were NT by GID only) and two NT isolates from both methods, and their <i>in silico</i> serotypes were obtained from genome sequencing based on their capsule loci. The mPCR results were supported by the <i>in silico</i> serotyping of the seven serotype-discrepant isolates. The discrepant results and NT isolates determined by mPCR were attributed to deletions and unknown sequences in the serotype-specific region of each capsule locus. Compared with previous investigations, this study found a similar predominant serotype profile, but a different prevalence frequency for <i>H</i>. <i>parasuis</i>, and the five most prevalent serotypes or strain groups were serotypes 5, 4, NT, 7 and 13 for mPCR, and serotypes 5, NT, 4, 7 and 13/10/14 for GID. Additionally, serotype 7 was recognized as a principal serotype in this work.</p></div

General genome features of the sequenced <i>H</i>. <i>parasuis</i> isolates.

<p>General genome features of the sequenced <i>H</i>. <i>parasuis</i> isolates.</p

Comparison of the results for mPCR and GID for 100 <i>H</i>. <i>parasuis</i> isolates.

<p>Comparison of the results for mPCR and GID for 100 <i>H</i>. <i>parasuis</i> isolates.</p