Biomarkers of intestinal barrier function in multiple sclerosis are associated with disease activity.

BACKGROUND: Recent evidence suggests a role for the gut-brain axis in the pathophysiology of multiple sclerosis (MS). MATERIALS AND METHODS: We studied biomarkers of intestinal permeability in 126 people with MS (57 relapsing-remitting multiple sclerosis (RRMS) and 69 progressive MS) and in a group of healthy controls for comparison. Serum/plasma concentrations of zonulin (a regulator of enterocyte tight junctions), tight junction proteins (ZO-1 and occludin), intestinal fatty acid binding protein (IFABP)/ileal bile acid binding protein (IBABP), D-lactate, and lipopolysaccharide (LPS) binding protein were measured. RESULTS: Zonulin concentrations were significantly higher when a concurrent magnetic resonance imaging (MRI) confirmed the presence of blood-brain barrier (BBB) disruption (Gad+ RRMS) and were correlated with tight junction proteins. IBABP and D-lactate were elevated in people with RRMS compared to controls, but did not discriminate between Gad+ and Gad- subgroups. Baseline zonulin concentrations were associated with 1-year disease progression in progressive MS. CONCLUSIONS: People with MS have altered biomarkers of intestinal barrier integrity. Zonulin concentrations are associated with 1-year disease progression in progressive MS and closely mirror BBB breakdown in RRMS. Zonulin may mediate breakdown of both the intestinal barrier and the BBB in gut dysbiosis through the regulation of tight junctions. This could explain how the gut-brain axis modulates neuroinflammation in MS.

Feasibility study to assess lesion repair in relapsing-remitting multiple sclerosis: A randomized controlled pilot clinical trial of domperidone add-on treatment.

BACKGROUND: Identification of therapies to promote repair in multiple sclerosis is challenged by the lack of an accepted trial model and associated outcome measures. The goal of this study was to determine the feasibility of a new trial model that enrolls disease modifying therapy (DMT)-treated relapsing-remitting multiple sclerosis (RRMS) participants who have enhancing lesions on clinically indicated brain MRI, and to explore estimates of lesion repair using MRI. METHODS: This was a single site randomized controlled clinical trial. Recruitment took place between November 2015 and January 2019, with final follow-up in February 2019. DMT-treated RRMS participants aged 18-60 years with at least one gadolinium-enhancing lesion on clinically indicated brain MRI were included. Participants were randomized 2:1 to oral domperidone add-on 10-mg three times daily for 16 weeks or no add-on treatment (control). The primary outcomes were feasibility of the model pre-defined as recruitment of 24 participants within 36 months with a 79 % completion rate, and MRI outcomes of lesion repair measured at 16 and 32 weeks using texture analysis, magnetization transfer imaging (MTI), and diffusion tensor imaging (DTI). The impact of domperidone on serum prolactin at 6 and 16 weeks was also evaluated. RESULTS: Of 237 RRMS participants screened, 17 (14 women) were randomized: 12 to domperidone add-on and 5 to control. All completed the study. Median (range) age was 38.9 (26.7-55.9) years; EDSS was 1.5 (1.0-3.5); and disease duration was 12.9 (2.9-23.3) years. Both groups showed improvement in MRI texture and diffusion fractional anisotropy (FA) at 32 weeks, and the domperidone group demonstrated additional recovery at 16 weeks in both texture and FA. There was no significant group difference in any MRI outcome. Of the 12 domperidone participants, 7 had ≥4x higher serum prolactin than normal. There were no serious adverse events. CONCLUSION: The recruitment target was not met and therefore the trial model was not feasible despite a full completion rate. The imaging techniques performed well, especially MRI texture analysis, suggesting the sample size being sufficient for estimating lesion repair. The main challenge of this trial model may be recruiting gadolinium-enhancing lesions in DMT-treated RRMS participants. Prolactin is safe and may hold promise as a remyelination therapy. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02493049.

Genomic comparison of Salmonella typhimurium DT104 with non-DT104 strains.

DT104 emerged as a new branch of Salmonella typhimurium with resistance to multiple antimicrobials. To reveal some general genomic features of DT104 for clues of evolutionary events possibly associated with the emergence of this relatively new type of this pathogen, we mapped 11 independent DT104 strains and compared them with non-DT104 S. typhimurium strains. We found that all 11 DT104 strains contained three insertions absent in non-DT104 strains, i.e., the previously reported ST104, ST104B and ST64B. However, SGI-1, a genomic island known to be responsible for DT104 multidrug resistance, was not present in all DT104 strains examined in this study: one DT104 strain did not contain SGI-1 but carried a 144 kb plasmid, suggesting possible evolutionary relationships between the two DNA elements in the development of antimicrobial resistance.

Distinct characteristics and severity of brain magnetic resonance imaging lesions in women and men with multiple sclerosis assessed using verified texture analysis measures.

Background and goal: In vivo characterization of brain lesion types in multiple sclerosis (MS) has been an ongoing challenge. Based on verified texture analysis measures from clinical magnetic resonance imaging (MRI), this study aimed to develop a method to identify two extremes of brain MS lesions that were approximately severely demyelinated (sDEM) and highly remyelinated (hREM), and compare them in terms of common clinical variables. Method: Texture analysis used an optimized gray-level co-occurrence matrix (GLCM) method based on FLAIR MRI from 200 relapsing-remitting MS participants. Two top-performing metrics were calculated: texture contrast and dissimilarity. Lesion identification applied a percentile approach according to texture values calculated: ≤ 25 percentile for hREM and ≥75 percentile for sDEM. Results: The sDEM had a greater total normalized volume yet smaller average size, and worse MRI texture than hREM. In lesion distribution mapping, the two lesion types appeared to overlap largely in location and were present the most in the corpus callosum and periventricular regions. Further, in sDEM, the normalized volume was greater and in hREM, the average size was smaller in men than women. There were no other significant results in clinical variable-associated analyses. Conclusion: Percentile statistics of competitive MRI texture measures may be a promising method for probing select types of brain MS lesion pathology. Associated findings can provide another useful dimension for improved measurement and monitoring of disease activity in MS. The different characteristics of sDEM and hREM between men and women likely adds new information to the literature, deserving further confirmation.

Advanced diffusion MRI and image texture analysis detect widespread brain structural differences between relapsing-remitting and secondary progressive multiple sclerosis.

Introduction: Disease development in multiple sclerosis (MS) causes dramatic structural changes, but the exact changing patterns are unclear. Our objective is to investigate the differences in brain structure locally and spatially between relapsing-remitting MS (RRMS) and its advanced form, secondary progressive MS (SPMS), through advanced analysis of diffusion magnetic resonance imaging (MRI) and image texture. Methods: A total of 20 patients with RRMS and nine patients with SPMS from two datasets underwent 3T anatomical and diffusion tensor imaging (DTI). The DTI was harmonized, augmented, and then modeled, which generated six voxel- and sub-voxel-scale measures. Texture analysis focused on T2 and FLAIR MRI, which produced two phase-based measures, namely, phase congruency and weighted mean phase. Data analysis was 3-fold, i.e., histogram analysis of whole-brain normal appearing white matter (NAWM); region of interest (ROI) analysis of NAWM and lesions within three critical white matter tracts, namely, corpus callosum, corticospinal tract, and optic radiation; and along-tract statistics. Furthermore, by calculating the z-score of core-rim pathology within lesions based on diffusion measures, we developed a novel method to define chronic active lesions and compared them between cohorts. Results: Histogram features from diffusion and all but one texture measure differentiated between RRMS and SPMS. Within-tract ROI analysis detected cohort differences in both NAWM and lesions of the corpus callosum body in three measures of neurite orientation and anisotropy. Along-tract statistics detected cohort differences from multiple measures, particularly lesion extent, which increased significantly in SPMS in posterior corpus callosum and optic radiations. The number of chronic active lesions were also significantly higher (by 5-20% over z-scores 0.5 and 1.0) in SPMS than RRMS based on diffusion anisotropy, neurite content, and diameter. Conclusion: Advanced diffusion MRI and texture analysis may be promising approaches for thorough understanding of brain structural changes from RRMS to SPMS, thereby providing new insight into disease development mechanisms in MS.

Advanced Analysis of Diffusion Tensor Imaging Along With Machine Learning Provides New Sensitive Measures of Tissue Pathology and Intra-Lesion Activity in Multiple Sclerosis.

Tissue pathology in multiple sclerosis (MS) is highly complex, requiring multi-dimensional analysis. In this study, our goal was to test the feasibility of obtaining high angular resolution diffusion imaging (HARDI) metrics through single-shell modeling of diffusion tensor imaging (DTI) data, and investigate how advanced measures from single-shell HARDI and DTI tractography perform relative to classical DTI metrics in assessing MS pathology. We examined 52 relapsing-remitting MS patients who had 3T anatomical brain MRI and DTI. Single-shell HARDI modeling yielded 5 sub-voxel-based metrics, totalling 11 diffusion measures including 4 DTI and 2 tractography metrics. Based on machine learning of 3-dimensional regions of interest, we evaluated the importance of the measures through several tissue classification tasks. These included two within-subject comparisons: lesion versus normal appearing white matter (NAWM); and lesion core versus shell. Further, by stratifying patients as having high (above 75% ile ) and low (below 25% ile ) number of MS lesions, we also performed 2 classifications between subjects for lesions and NAWM respectively. Results showed that in lesion-NAWM analysis, HARDI orientation distribution function (ODF) energy, DTI fractional anisotropy (FA), and HARDI orientation dispersion index were the top three metrics, which together achieved 65.2% accuracy and 0.71 area under the receiver operating characteristic curve (AUROC). In core-shell analysis, DTI mean diffusivity (MD), radial diffusivity, and FA were the top three metrics, and MD dominated the classification, which achieved 59.3% accuracy and 0.59 AUROC alone. Between patients, FA was the leading feature in lesion comparisons, while ODF energy was the best in NAWM separation. Collectively, single-shell modeling of common diffusion data can provide robust orientation measures of lesion and NAWM pathology, and DTI metrics are most sensitive to intra-lesion abnormality. Combined analysis of both advanced and classical diffusion measures may be critical for improved understanding of MS pathology.

Multiple genetic switches spontaneously modulating bacterial mutability.

BACKGROUND: All life forms need both high genetic stability to survive as species and a degree of mutability to evolve for adaptation, but little is known about how the organisms balance the two seemingly conflicting aspects of life: genetic stability and mutability. The DNA mismatch repair (MMR) system is essential for maintaining genetic stability and defects in MMR lead to high mutability. Evolution is driven by genetic novelty, such as point mutation and lateral gene transfer, both of which require genetic mutability. However, normally a functional MMR system would strongly inhibit such genomic changes. Our previous work indicated that MMR gene allele conversion between functional and non-functional states through copy number changes of small tandem repeats could occur spontaneously via slipped-strand mis-pairing during DNA replication and therefore may play a role of genetic switches to modulate the bacterial mutability at the population level. The open question was: when the conversion from functional to defective MMR is prohibited, will bacteria still be able to evolve by accepting laterally transferred DNA or accumulating mutations? RESULTS: To prohibit allele conversion, we "locked" the MMR genes through nucleotide replacements. We then scored changes in bacterial mutability and found that Salmonella strains with MMR locked at the functional state had significantly decreased mutability. To determine the generalizability of this kind of mutability 'switching' among a wider range of bacteria, we examined the distribution of tandem repeats within MMR genes in over 100 bacterial species and found that multiple genetic switches might exist in these bacteria and may spontaneously modulate bacterial mutability during evolution. CONCLUSIONS: MMR allele conversion through repeats-mediated slipped-strand mis-pairing may function as a spontaneous mechanism to switch between high genetic stability and mutability during bacterial evolution.

Gadolinium enhancement on cranial MRI in multiple sclerosis is age dependent.

BACKGROUND: Epidemiological, pathological and radiological studies suggest that inflammatory demyelination in MS is an age-dependent process, and that the formation of focal inflammatory demyelinating lesions decreases with age. Gadolinium-enhancing lesions are a biomarker of inflammatory disease activity in MS, but little is known about the relation of age and gadolinium enhancement on cranial MRI scans in people with MS. In this study, we investigated the association of age and other risk factors with gadolinium enhancement on cranial MRI in a retrospective cross-sectional clinical MS cohort. METHODS: In a cohort including 1543 people with CIS and MS, we investigated the association of the risk factors age, sex, disease course, immunomodulatory drug (IMD) treatment, and disability with gadolinium enhancement on cranial MRI scans using a binary logistic regression model. RESULTS: Age was the most important factor associated with gadolinium enhancement, with the odds of gadolinium enhancement decreasing with advancing age. Participants with CIS had lower odds of gadolinium enhancement (odds ratio of 0.42, 95% confidence interval of 0.24-0.72 compared to RRMS). Sex, disease course and IMD treatment were not significantly associated with gadolinium enhancement in our cohort. CONCLUSIONS: Our investigation shows that gadolinium enhancement is strongly associated with age. Since gadolinium enhancement is a marker of inflammatory disease activity, our findings suggest that inflammatory disease activity declines with age, and that IMD treatment may be more beneficial in younger and less useful in older people with MS.

E. coli diversity: low in colorectal cancer.

BACKGROUND: Escherichia coli are mostly commensals but also contain pathogenic lineages. It is largely unclear whether the commensal E. coli as the potential origins of pathogenic lineages may consist of monophyletic or polyphyletic populations, elucidation of which is expected to lead to novel insights into the associations of E. coli diversity with human health and diseases. METHODS: Using genomic sequencing and pulsed field gel electrophoresis (PFGE) techniques, we analyzed E. coli from the intestinal microbiota of three groups of healthy individuals, including preschool children, university students, and seniors of a longevity village, as well as colorectal cancer (CRC) patients, to probe the commensal E. coli populations for their diversity. RESULTS: We delineated the 2280 fresh E. coli isolates from 185 subjects into distinct genome types (genotypes) by PFGE. The genomic diversity of the sampled E. coli populations was so high that a given subject may have multiple genotypes of E. coli, with the general diversity within a host going up from preschool children through university students to seniors. Compared to the healthy subjects, the CRC patients had the lowest diversity level among their E. coli isolates. Notably, E. coli isolates from CRC patients could suppress the growth of E. coli bacteria isolated from healthy controls under nutrient-limited culture conditions. CONCLUSIONS: The coexistence of multiple E. coli lineages in a host may help create and maintain a microbial environment that is beneficial to the host. As such, the low diversity of E. coli bacteria may be associated with unhealthy microenvironment in the intestine and hence facilitate the pathogenesis of diseases such as CRC.

Locally synthesized calcitonin gene-related Peptide has a critical role in peripheral nerve regeneration.

Regeneration of peripheral nerves involves complex and intimate interactions between axons and Schwann cells. Here, we show that local axon synthesis and action of the neuropeptide calcitonin gene-related peptide (CGRP) is critical for this collaboration. After peripheral sural sensory axon injury in rats, we observed an unexpectedly large proportion of axons that newly expressed CGRP during regeneration. Intense peptide expression accompanied local rises in alphaCGRP mRNA in the nerve trunk, and there was evidence of transport of alphaCGRP mRNA into regenerating axons, indicating intra-axonal peptide synthesis. Calcitonin gene-related peptide receptor and its receptor activity modifying protein were expressed onadjacent Schwann cells, where they were available for signaling. Moreover, exogenous CGRP induced proliferation in isolated adult Schwann cells. New axon outgrowth and CGRP expression depended on local peptide synthesis and were inhibited by exposure tolocal translation inhibitors. Local delivery of siRNAs to either alphaCGRP or receptor activity modifying protein 1 to sites of nerve transection was associated with severe disruption of axon outgrowth.These findings indicate that robust localized intra-axonal translation of the CGRP neuropeptide during regeneration signals Schwann cell proliferation, behavior that is critical for partnering during adult peripheral nerve regrowth.

Sudden infant death syndrome (SIDS) in African Americans: polymorphisms in the gene encoding the stress peptide pituitary adenylate cyclase-activating polypeptide (PACAP).

AIMS: Mice lacking pituitary adenylate cyclase-activating polypeptide (PACAP) are prone to sudden death in the second post-natal week, having respiratory and metabolic disturbances reminiscent of the human Sudden Infant Death Syndrome (SIDS). Here we test the hypothesis that the human PACAP gene is a site of genetic variance associated with SIDS in a cohort of 92 victims and 92 matched controls. METHODS: Using polymerase chain reaction and sequencing, we examined the PACAP gene in 92 SIDS cases (46 Caucasians and 46 African Americans) and 92 race- and gender-matched controls. RESULTS: We found no significant associations between PACAP and SIDS in Caucasians. However, in the African Americans, a non-synonymous single nucleotide polymorphism (i.e. an aspartic acid/glycine coding variant, rs2856966) within exon 2 of PACAP was significantly associated with SIDS (p = 0.004), as were haplotypes containing this polymorphism (p < 0.0001). Glycine was three times more likely at this location in the African-American SIDS victims (17 cases) than African-American controls (5 cases). CONCLUSION: These data are the first to suggest an association between a variant within the coding region of the PACAP gene and SIDS. Based on these findings, further investigations are warranted into the functional importance of PACAP signaling in neonatal survival and the role of PACAP-signaling abnormalities in SIDS.

Domperidone-induced elevation of serum prolactin levels and immune response in multiple sclerosis.

Increasing systemic prolactin levels improves remyelination and neuronal survival in animal models of Multiple Sclerosis (MS), but it has been suggested that this therapeutic strategy may also increase inflammatory responses, and potentially harm patients. We analyzed serum prolactin and cytokine, chemokine and growth factor levels in sera from MS patients enrolled in two clinical trials who were treated with domperidone, a generic drug that increases systemic prolactin levels. In patients treated with domperidone, molecule levels changed little during follow up, while prolactin levels increased several-fold. We found no significant association between prolactin levels and radiological or clinical outcome.

Diverse genome structures of Salmonella paratyphi C.

BACKGROUND: Salmonella paratyphi C, like S. typhi, is adapted to humans and causes typhoid fever. Previously we reported different genome structures between two strains of S. paratyphi C, which suggests that S. paratyphi C might have a plastic genome (large DNA segments being organized in different orders or orientations on the genome). As many but not all host-adapted Salmonella pathogens have large genomic insertions as well as the supposedly resultant genomic rearrangements, bacterial genome plasticity presents an extraordinary evolutionary phenomenon. Events contributing to genomic plasticity, especially large insertions, may be associated with the formation of particular Salmonella pathogens. RESULTS: We constructed a high resolution genome map in S. paratyphi C strain RKS4594 and located four insertions totaling 176 kb (including the 90 kb SPI7) and seven deletions totaling 165 kb relative to S. typhimurium LT2. Two rearrangements were revealed, including an inversion of 1602 kb covering the ter region and the translocation of the 43 kb I-CeuI F fragment. The 23 wild type strains analyzed in this study exhibited diverse genome structures, mostly as a result of recombination between rrn genes. In at least two cases, the rearrangements involved recombination between genomic sites other than the rrn genes, possibly homologous genes in prophages. Two strains had a 20 kb deletion between rrlA and rrlB, which is a highly conservative region and no deletion has been reported in this region in any other Salmonella lineages. CONCLUSION: S. paratyphi C has diverse genome structures among different isolates, possibly as a result of large genomic insertions, e.g., SPI7. Although the Salmonella typhoid agents may not be more closely related among them than each of them to other Salmonella lineages, they may have evolved in similar ways, i.e., acquiring typhoid-associated genes followed by genome structure rearrangements. Comparison of multiple Salmonella typhoid agents at both single sequenced genome and population levels will facilitate the studies on the evolutionary process of typhoid pathogenesis, especially the identification of typhoid-associated genes.

Remyelination therapies: a new direction and challenge in multiple sclerosis.

Multiple sclerosis is characterized by inflammatory activity that results in destruction of the myelin sheaths that enwrap axons. The currently available medications for multiple sclerosis are predominantly immune-modulating and do not directly promote repair. White matter regeneration, or remyelination, is a new and exciting potential approach to treating multiple sclerosis, as remyelination repairs the damaged regions of the central nervous system. A wealth of new strategies in animal models that promote remyelination, including the repopulation of oligodendrocytes that produce myelin, has led to several clinical trials to test new reparative therapies. In this Review, we highlight the biology of, and obstacles to, remyelination. We address new strategies to improve remyelination in preclinical models, highlight the therapies that are currently undergoing clinical trials and discuss the challenges of objectively measuring remyelination in trials of repair in multiple sclerosis.

Differential degeneration of the ACTAGT sequence among Salmonella: a reflection of distinct nucleotide amelioration patterns during bacterial divergence.

When bacteria diverge, they need to adapt to the new environments, such as new hosts or different tissues of the same host, by accumulating beneficial genomic variations, but a general scenario is unknown due to the lack of appropriate methods. Here we profiled the ACTAGT sequence and its degenerated forms (i.e., hexa-nucleotide sequences with one of the six nucleotides different from ACTAGT) in Salmonella to estimate the nucleotide amelioration processes of bacterial genomes. ACTAGT was mostly located in coding sequences but was also found in several intergenic regions, with its degenerated forms widely scattered throughout the bacterial genomes. We speculated that the distribution of ACTAGT and its degenerated forms might be lineage-specific as a consequence of different selection pressures imposed on ACTAGT at different genomic locations (in genes or intergenic regions) among different Salmonella lineages. To validate this speculation, we modelled the secondary structures of the ACTAGT-containing sequences conserved across Salmonella and many other enteric bacteria. Compared to ACTAGT at conserved regions, the degenerated forms were distributed throughout the bacterial genomes, with the degeneration patterns being highly similar among bacteria of the same phylogenetic lineage but radically different across different lineages. This finding demonstrates biased amelioration under distinct selection pressures among the bacteria and provides insights into genomic evolution during bacterial divergence.

CTAG-containing cleavage site profiling to delineate Salmonella into natural clusters.

BACKGROUND: The bacterial genus Salmonella contains thousands of serotypes that infect humans or other hosts, causing mild gastroenteritis to potentially fatal systemic infections in humans. Pathogenically distinct Salmonella serotypes have been classified as individual species or as serological variants of merely one or two species, causing considerable confusion in both research and clinical settings. This situation reflects a long unanswered question regarding whether the Salmonella serotypes exist as discrete genetic clusters (natural species) of organisms or as phenotypic (e.g. pathogenic) variants of a single (or two) natural species with a continuous spectrum of genetic divergence among them. Our recent work, based on genomic sequence divergence analysis, has demonstrated that genetic boundaries exist among Salmonella serotypes, circumscribing them into clear-cut genetic clusters of bacteria. METHODOLOGIES/PRINCIPAL FINDINGS: To further test the genetic boundary concept for delineating Salmonella into clearly defined natural lineages (e.g., species), we sampled a small subset of conserved genomic DNA sequences, i.e., the endonuclease cleavage sites that contain the highly conserved CTAG sequence such as TCTAGA for XbaI. We found that the CTAG-containing cleavage sequence profiles could be used to resolve the genetic boundaries as reliably and efficiently as whole genome sequence comparisons but with enormously reduced requirements for time and resources. CONCLUSIONS: Profiling of CTAG sequence subsets reflects genetic boundaries among Salmonella lineages and can delineate these bacteria into discrete natural clusters.

mutL as a genetic switch of bacterial mutability: turned on or off through repeat copy number changes.

Bacterial adaptation to changing environments can be achieved through the acquisition of genetic novelty by accumulation of mutations and recombination of laterally transferred genes into the genome, but the mismatch repair (MMR) system strongly inhibits both these types of genetic changes. As mutation and recombination do occur in bacteria, it is of interest to understand how genetic novelty may be achieved in the presence of MMR. Previously, we observed associations of a defective MMR genotype, 6bpΔmutL, with greatly elevated bacterial mutability in Salmonella typhimurium. To validate these observations, we experimentally converted the mutL gene between the wild-type and 6bpΔmutL in S. typhimurium and inspected the bacterial mutability status. When 6bpΔmutL was converted to mutL, the originally highly mutable Salmonella strains regained genetic stability; when mutL was converted to 6bpΔmutL, the mutability was elevated 100-fold. Interestingly, mutL cells were found to grow out of 6bpΔmutL cells; the new mutL cells eventually replaced the original 6bpΔmutL population. As conversion between mutL and 6bpΔmutL may occur readily during DNA replication, it may represent a previously unrecognized mechanism to modulate bacterial mutability at the population level, allowing bacteria to respond rapidly to changing environments while minimizing the risks associated with persistent hypermutability.

Salmonella paratyphi C: genetic divergence from Salmonella choleraesuis and pathogenic convergence with Salmonella typhi.

BACKGROUND: Although over 1400 Salmonella serovars cause usually self-limited gastroenteritis in humans, a few, e.g., Salmonella typhi and S. paratyphi C, cause typhoid, a potentially fatal systemic infection. It is not known whether the typhoid agents have evolved from a common ancestor (by divergent processes) or acquired similar pathogenic traits independently (by convergent processes). Comparison of different typhoid agents with non-typhoidal Salmonella lineages will provide excellent models for studies on how similar pathogens might have evolved. METHODOLOGIES/PRINCIPAL FINDINGS: We sequenced a strain of S. paratyphi C, RKS4594, and compared it with previously sequenced Salmonella strains. RKS4594 contains a chromosome of 4,833,080 bp and a plasmid of 55,414 bp. We predicted 4,640 intact coding sequences (4,578 in the chromosome and 62 in the plasmid) and 152 pseudogenes (149 in the chromosome and 3 in the plasmid). RKS4594 shares as many as 4346 of the 4,640 genes with a strain of S. choleraesuis, which is primarily a swine pathogen, but only 4008 genes with another human-adapted typhoid agent, S. typhi. Comparison of 3691 genes shared by all six sequenced Salmonella strains placed S. paratyphi C and S. choleraesuis together at one end, and S. typhi at the opposite end, of the phylogenetic tree, demonstrating separate ancestries of the human-adapted typhoid agents. S. paratyphi C seemed to have suffered enormous selection pressures during its adaptation to man as suggested by the differential nucleotide substitutions and different sets of pseudogenes, between S. paratyphi C and S. choleraesuis. CONCLUSIONS: S. paratyphi C does not share a common ancestor with other human-adapted typhoid agents, supporting the convergent evolution model of the typhoid agents. S. paratyphi C has diverged from a common ancestor with S. choleraesuis by accumulating genomic novelty during adaptation to man.