Search

To understand the fine differential elements that can lead to or prevent acute respiratory distress syndrome (ARDS) in COVID-19 patients, it is crucial to investigate the immune response architecture. We herein dissected the multiple layers of B cell responses by flow cytometry and Ig repertoire analysis from acute phase to recovery. Flow cytometry with FlowSOM analysis showed major changes associated with COVID-19 inflammation such as an increase of double-negative B-cells and ongoing plasma cell differentiation. This paralleled COVID-19-driven expansion of two disconnected B-cell repertoires. Demultiplexing successive DNA and RNA Ig repertoire patterns characterized an early expansion of IgG1 clonotypes with atypically long and uncharged CDR3, the abundance of this inflammatory repertoire being correlated with ARDS and likely pejorative. A superimposed convergent response included convergent anti-SARS-CoV-2 clonotypes. It featured progressively increasing somatic hypermutation together with normal-length or short CDR3 and it persisted until a quiescent memory B-cell stage after recovery.

Mean values for irregularity index and relative lesion area of Phytophthora ramorum isolates identified in K-means analysis. Isolates in Cluster 3 were significantly different from those in Clusters 1 and 2. Bars with different letters are significantly different at pÂ

Allele sizes for the 15 multilocus genotypes (MLGs) identified in 30 Washington State isolates of P. ramorum. (PDF 48 kb)

The maximum and minimum radii within a 45-degree sector of the colony were used to obtain irregularity index as: (43.5â 34.7)/43.5â =â 20.8% (PDF 960 kb)

Diverse CCNVs revealed by BIC-seq analysis (upper graph for each panel) and a read-depth analysis for heterozygous allele ratios using 10 Kb long non-overlapping sliding window (lower graph). A concatenated view of the 30 largest scaffolds with the total length of 30Â MB, which corresponding to approximately a half of the total genome of Phytophthora ramorum, are shown. Scaffolds numbers for large CCNV regions are indicated with pink bars, and those for copy number neutral LOH are shown with blue bars. Scales show log (base 2) fold difference between sample isolates and reference isolates for BIC-seq analysis and log (base 2) ratios of alleles of sample isolates for the heterozygous allele ratio analysis. At each heterozygous locus, a read count ratio (more-abundant allele/less-abundant allele) was calculated. A) A re-isolate 9D1 from a log of coast live oak showing a small copy number change at scaffold 9 in comparison to its original isolate Pr-1556 (wt, bay) used as a reference. 9D1 is likely a heterokaryon (HK) carrying trisomic (3Ă CCNV) and euploid nuclei. B) Pr-16 carries trisomy and cnLOH aberrations. C) MK516a carries cnLOH. D) BS2014â 584 shows a complicated patter indicating it carries a mixture of trisomic and monosomic nuclei. E) Pr-140.7 is a complicated monosomic heterokaryon. F) Pr-140.9 is monosomic at scaffold 9. G) Pr-106 is a normal euploid. H) Pr-486 is a trisomy at scaffold 10. Numbers of short segmental CNVs are also seen. I) and J) Both re-isolates Pr-745#4 and Pr-1556#7#1 carry complicated mixtures of monosomic and trisomic nuclei. K) BS96 from California bay is trisomic at scaffold 9. L) Pr-218 from Rhamnus cathartica is trisomic at scaffold 10. M) Pr-102, the sequence isolate (Tyler et al..., [17]) is trisomic at scaffold 10. N) a re-isolate Pr-745#3 is trisomic. O) Pr-455 from Osmorhiza berteroi shows cnLOH at several scaffolds. P) Pr-472 from Choisya ternate is trisomic. (PDF 3465 kb)

Colony morphology of Washington State isolates seen on Petri plates. Nwt phenotype is indicated. Colonies were grown on solid 1× CV8A medium for 7 days at 20 °C in dark. WSU115–0077 initially showed wt colony morphology, however, it displayed nwt in subsequent subculture. (PDF 1200 kb)

Diverse CCNVs revealed by BIC-seq analysis (upper graph for each panel) and a read-depth analysis for heterozygous allele ratios using 10 Kb long non-overlapping sliding window (lower graph). A concatenated view of the 30 largest scaffolds with the total length of 30Â MB, which corresponding to approximately a half of the total genome of Phytophthora ramorum, are shown. Scaffolds numbers for large CCNV regions are indicated with pink bars, and those for copy number neutral LOH are shown with blue bars. Scales show log (base 2) fold difference between sample isolates and reference isolates for BIC-seq analysis and log (base 2) ratios of alleles of sample isolates for the heterozygous allele ratio analysis. At each heterozygous locus, a read count ratio (more-abundant allele/less-abundant allele) was calculated. A) A re-isolate 9D1 from a log of coast live oak showing a small copy number change at scaffold 9 in comparison to its original isolate Pr-1556 (wt, bay) used as a reference. 9D1 is likely a heterokaryon (HK) carrying trisomic (3Ă CCNV) and euploid nuclei. B) Pr-16 carries trisomy and cnLOH aberrations. C) MK516a carries cnLOH. D) BS2014â 584 shows a complicated patter indicating it carries a mixture of trisomic and monosomic nuclei. E) Pr-140.7 is a complicated monosomic heterokaryon. F) Pr-140.9 is monosomic at scaffold 9. G) Pr-106 is a normal euploid. H) Pr-486 is a trisomy at scaffold 10. Numbers of short segmental CNVs are also seen. I) and J) Both re-isolates Pr-745#4 and Pr-1556#7#1 carry complicated mixtures of monosomic and trisomic nuclei. K) BS96 from California bay is trisomic at scaffold 9. L) Pr-218 from Rhamnus cathartica is trisomic at scaffold 10. M) Pr-102, the sequence isolate (Tyler et al..., [17]) is trisomic at scaffold 10. N) a re-isolate Pr-745#3 is trisomic. O) Pr-455 from Osmorhiza berteroi shows cnLOH at several scaffolds. P) Pr-472 from Choisya ternate is trisomic. (PDF 3465 kb)

Diverse CCNVs revealed by BIC-seq analysis (upper graph for each panel) and a read-depth analysis for heterozygous allele ratios using 10 Kb long non-overlapping sliding window (lower graph). A concatenated view of the 52 largest scaffolds with the total length of 300 MB, which corresponding to approximately a half of the total genome of Phytophthora ramorum, are shown. Scaffolds numbers for large CCNV regions are indicated with pink bars, and those for LOH are shown with green bars. Scales show log (base 2) fold difference between sample isolates and reference isolates for BIC-seq analysis and log (base 2) ratios of alleles of sample isolates for the heterozygous allele ratio analysis. At each heterozygous locus, a read count ratio (more-abundant allele/less-abundant allele) was calculated. A) Pr-102, the genome sequence isolate is trisomic. B) Oak isolate Pr-16 is trisomic as well as cnLOH. C) Trisomy persists in a re-isolate of Pr-102 from California bay. D) A re-isolate Pr745#4 and E) an oak isolate Pr-140.7 are CCNV heterokaryons. F) nwt EU1 isolate P2386 revealed CCNVs and LOH when wt EU1 isolate P2363 was used as a reference. (PDF 1014 kb)

BIC-seq analysis for three EU1 isolates using an NA1 isolate Pr-1556 (wt, bay) as a reference. Due to the genome divergence between the EU1 and NA1 lineages, a high background was seen. For an explanation of graphs, see Additional file 3. A) When the genome contents of EU1 wt isolate P2363 and the NA1 reference were compared, copy number reduction was detected for several regions (red bars), which accompany increase in SNP allele ratios, suggesting P2363 is a mixture of monosomic and disomic nuclei. B) When EU1 nwt isolate P2346 and the reference were compared, increases (red bars) as well as decreases in the genome content were detected. Note that several scaffolds show large heterozygous SNP ratios while copy number changes in corresponding scaffolds are subtle. This may implicate a formation of cnLOH. C) EU1 nwt isolate P2386 shows large SNP allele ratios in several scaffolds indicating the formation of cnLOH (red bars). These cnLOH scaffolds are, however, distinct from those observed in P2346. (PDF 561 kb)

The chromosomal breakpoint in scaffold 37. From the right most heterozygous SNP and the left most SNP with LOH, the breakpoint for Pr-16 (cnLOH) was inferred to be within the 3.0kb red rectangle region (top panel, Integrative Genomic Viewer ver. 2.3.34). Note that Pr-1556 is a normal diploid isolate having heterozygous SNP loci on the right of the breakpoint. Homologous chromosomes are depicted in orange and cyan (lower panel). (PDF 51 kb)

The chromosomal breakpoint in scaffold 44. From the right most heterozygous SNP and the left most SNP with LOH, the breakpoint for Pr-16 (cnLOH) was inferred to be within the 1.3kb red rectangle region (top panel, Integrative Genomic Viewer ver. 2.3.34). Homologous chromosomes are depicted in orange and cyan (lower panel). (PDF 46 kb)

Nwt colony morphology seen among EU1 isolates obtained from a single 4 m long lesion on a mature Lawson cypress. EU1 isolates (A-C) are derived from the top, (D-F) are from the middle, and (G-I) are from the bottom of the lesion. Mnwt and snwt indicate moderate and severe non-wild types, respectively. (PDF 234 kb)

The chromosomal breakpoint in scaffold 34. From the right end of the 5.0 kb indel and the left most heterozygous SNP, the breakpoint for MK516a (cnLOH) is inferred to be somewhere within the 7.5 kb red rectangle region (top panel, Integrative Genomics Viewer ver. 2.3.34). For Pr-102 (trisomy) and Pr-16 (trisomy), ratios of heterozygous SNP reads are not reliable for inference for the precise transition from disomy to trisomy. BICseq CCNV analysis, however, located the breakpoints for Pr-102 and Pr-16 at positions 191,470 bp and 190,370 bp, respectively (only Pr-16 is shown in the middle panel), which are within the red rectangle range. Homologous chromosomes are depicted in orange and cyan (lower panel). (PDF 63 kb)

The chromosomal breakpoint in scaffold 12. From the right most heterozygous SNP and the left most SNP with LOH, the breakpoints for Pr-140.9 (monosomy) and Pr-16 (cnLOH) were inferred to be within the 1.3kb red rectangle region (top panel, Integrative Genomic Viewer ver. 2.3.34). BICseq CCNV analysis for Pr-140.9 inferred the breakpoint in the close proximity to the red rectangle (middle panel). Homologous chromosomes are depicted in orange and cyan (lower panel). (PDF 57 kb)