Vivek Kapur

Interferon-inducible gene expression signature in peripheral blood cells of patients with severe lupus

Systemic lupus erythematosus (SLE) is a complex, inflammatory autoimmune disease that affects multiple organ systems. We used global gene expression profiling of peripheral blood mononuclear cells to identify distinct patterns of gene expression that distinguish most SLE patients from healthy controls. Strikingly, about half of the patients studied showed dysregulated expression of genes in the IFN pathway. Furthermore, this IFN gene expression “signature” served as a marker for more severe disease involving the kidneys, hematopoetic cells, and/or the central nervous system. These results provide insights into the genetic pathways underlying SLE, and identify a subgroup of patients who may benefit from therapies targeting the IFN pathway.

Managing Incidental Findings in Human Subjects Research: Analysis and Recommendations

No consensus yet exists on how to handle incidental findings (IFs) in human subjects research. Yet empirical studies document IFs in a wide range of research studies, where IFs are findings beyond the aims of the study that are of potential health or reproductive importance to the individual research participant. This paper reports recommendations of a two-year project group funded by NIH to study how to manage IFs in genetic and genomic research, as well as imaging research. We conclude that researchers have an obligation to address the possibility of discovering IFs in their protocol and communications with the IRB, and in their consent forms and communications with research participants. Researchers should establish a pathway for handling IFs and communicate that to the IRB and research participants. We recommend a pathway and categorize IFs into those that must be disclosed to research participants, those that may be disclosed, and those that should not be disclosed.

Genome sequence and comparative microarray analysis of serotype M18 group A Streptococcus strains associated with acute rheumatic fever outbreaks

Acute rheumatic fever (ARF), a sequelae of group A Streptococcus (GAS) infection, is the most common cause of preventable childhood heart disease worldwide. The molecular basis of ARF and the subsequent rheumatic heart disease are poorly understood. Serotype M18 GAS strains have been associated for decades with ARF outbreaks in the U.S. As a first step toward gaining new insight into ARF pathogenesis, we sequenced the genome of strain MGAS8232, a serotype M18 organism isolated from a patient with ARF. The genome is a circular chromosome of 1,895,017 bp, and it shares 1.7 Mb of closely related genetic material with strain SF370 (a sequenced serotype M1 strain). Strain MGAS8232 has 178 ORFs absent in SF370. Phages, phage-like elements, and insertion sequences are the major sources of variation between the genomes. The genomes of strain MGAS8232 and SF370 encode many of the same proven or putative virulence factors. Importantly, strain MGAS8232 has genes encoding many additional secreted proteins involved in human–GAS interactions, including streptococcal pyrogenic exotoxin A (scarlet fever toxin) and two uncharacterized pyrogenic exotoxin homologues, all phage-associated. DNA microarray analysis of 36 serotype M18 strains from diverse localities showed that most regions of variation were phages or phage-like elements. Two epidemics of ARF occurring 12 years apart in Salt Lake City, UT, were caused by serotype M18 strains that were genetically identical, or nearly so. Our analysis provides a critical foundation for accelerated research into ARF pathogenesis and a molecular framework to study the plasticity of GAS genomes.

The genome of Cryptosporidium hominis

Cryptosporidium species cause acute gastroenteritis and diarrhoea worldwide. They are members of the Apicomplexa—protozoan pathogens that invade host cells by using a specialized apical complex and are usually transmitted by an invertebrate vector or intermediate host. In contrast to other Apicomplexans, Cryptosporidium is transmitted by ingestion of oocysts and completes its life cycle in a single host. No therapy is available, and control focuses on eliminating oocysts in water supplies. Two species, C. hominis and C. parvum, which differ in host range, genotype and pathogenicity, are most relevant to humans. C. hominis is restricted to humans, whereas C. parvum also infects other mammals. Here we describe the eight-chromosome ∼9.2-million-base genome of C. hominis. The complement of C. hominis protein-coding genes shows a striking concordance with the requirements imposed by the environmental niches the parasite inhabits. Energy metabolism is largely from glycolysis. Both aerobic and anaerobic metabolisms are available, the former requiring an alternative electron transport system in a simplified mitochondrion. Biosynthesis capabilities are limited, explaining an extensive array of transporters. Evidence of an apicoplast is absent, but genes associated with apical complex organelles are present. C. hominis and C. parvum exhibit very similar gene complements, and phenotypic differences between these parasites must be due to subtle sequence divergence.

Complete genomic sequence of Pasteurella multocida,Pm70

We present here the complete genome sequence of a common avian clone of Pasteurella multocida, Pm70. The genome of Pm70 is a single circular chromosome 2,257,487 base pairs in length and contains 2,014 predicted coding regions, 6 ribosomal RNA operons, and 57 tRNAs. Genome-scale evolutionary analyses based on pairwise comparisons of 1,197 orthologous sequences between P. multocida, Haemophilus influenzae, and Escherichia coli suggest that P. multocida and H. influenzae diverged ≈270 million years ago and the γ subdivision of the proteobacteria radiated about 680 million years ago. Two previously undescribed open reading frames, accounting for ≈1% of the genome, encode large proteins with homology to the virulence-associated filamentous hemagglutinin of Bordetella pertussis. Consistent with the critical role of iron in the survival of many microbial pathogens, in silico and whole-genome microarray analyses identified more than 50 Pm70 genes with a potential role in iron acquisition and metabolism. Overall, the complete genomic sequence and preliminary functional analyses provide a foundation for future research into the mechanisms of pathogenesis and host specificity of this important multispecies pathogen.

Completion of the Genome Sequence of Brucella abortus and Comparison to the Highly Similar Genomes of Brucella melitensis and Brucella suis

Brucellosis is a worldwide disease of humans and livestock that is caused by a number of very closely related classical Brucella species in the alpha-2 subdivision of the Proteobacteria. We report the complete genome sequence of Brucella abortus field isolate 9-941 and compare it to those of Brucella suis 1330 and Brucella melitensis 16 M. The genomes of these Brucella species are strikingly similar, with nearly identical genetic content and gene organization. However, a number of insertion-deletion events and several polymorphic regions encoding putative outer membrane proteins were identified among the genomes. Several fragments previously identified as unique to either B. suis or B. melitensis were present in the B. abortus genome. Even though several fragments were shared between only B. abortus and B. suis, B. abortus shared more fragments and had fewer nucleotide polymorphisms with B. melitensis than B. suis. The complete genomic sequence of B. abortus provides an important resource for further investigations into determinants of the pathogenicity and virulence phenotypes of these bacteria.

GENETIC VARIATION IN PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS ISOLATES IN THE MIDWESTERN UNITED STATES

The nucleotide sequence of a 3266 bp region encompassing open reading frames (ORFs) 2 through 7 of the porcine reproductive and respiratory syndrome virus (PRRSV) was determined for 10 isolates recovered from the midwestern United States. Pairwise comparisons showed that genetic distances between isolates ranged from 2.5% to 7.9% (mean 5.8% +/- 0.2%) whereas the Lelystad strain from Europe was, on average, 34.8% divergent from US clones. Thus, US and European PRRSV isolates represent genetically distinct clusters of the same virus. ORF 5, which encodes the envelope glycoprotein, was the most polymorphic [total nucleotide diversity (pi) = 0.097 +/- 0.007] and ORF 6, encoding the viral M protein, was the most conserved (pi = 0.038 +/- 0.003). The substantial differences in nucleotide diversity among ORFs suggests that the virus is evolving by processes other than simple accumulation of random neutral mutations. In support of this hypothesis, statistical analyses of the nucleotide sequence provided strong evidence for intragenic recombination or gene conversion in ORFs 2, 3, 4, 5 and 7, but not in ORF 6. An excess of synonymous (silent) substitutions was observed in all six ORFs, indicating an evolutionary pressure to conserve amino acid sequences. Taken together, the data indicate that despite intragenic recombination among extant PRRSV isolates, purifying selection has acted to maintain the primary structure of individual ORFs.

Molecular Correlates of Host Specialization in Staphylococcus aureus

Background The majority of Staphylococcus aureus isolates that are recovered from either serious infections in humans or from mastitis in cattle represent genetically distinct sets of clonal groups. Moreover, population genetic analyses have provided strong evidence of host specialization among S. aureus clonal groups associated with human and ruminant infection. However, the molecular basis of host specialization in S. aureus is not understood. Methodology/Principal Findings We sequenced the genome of strain ET3-1, a representative isolate of a common bovine mastitis-causing S. aureus clone. Strain ET3-1 encodes several genomic elements that have not been previously identified in S. aureus, including homologs of virulence factors from other Gram-positive pathogens. Relative to the other sequenced S. aureus associated with human infection, allelic variation in ET3-1 was high among virulence and surface-associated genes involved in host colonization, toxin production, iron metabolism, antibiotic resistance, and gene regulation. Interestingly, a number of well-characterized S. aureus virulence factors, including protein A and clumping factor A, exist as pseudogenes in ET3-1. Whole-genome DNA microarray hybridization revealed considerable similarity in the gene content of highly successful S. aureus clones associated with bovine mastitis, but not among those clones that are only infrequently recovered from bovine hosts. Conclusions/Significance Whole genome sequencing and comparative genomic analyses revealed a set of molecular genetic features that distinguish clones of highly successful bovine-associated S. aureus optimized for mastitis pathogenesis in cattle from those that infect human hosts or are only infrequently recovered from bovine sources. Further, the results suggest that modern bovine specialist clones diverged from a common ancestor resembling human-associated S. aureus clones through a combination of foreign DNA acquisition and gene decay.

The evidence for Mycobacterium paratuberculosis in Crohn's disease.

Purpose of review Though long hypothesized, the putative link between Mycobacterium avium paratuberculosis and Crohns disease remains neither confirmed nor refuted. This article reviews published contributions that directly or indirectly address this question. Recent findings Epidemiologic studies, looking for M. avium paratuberculosis DNA in Crohns tissue, show a strong association between the agent and this disease. Supporting data, however, are presently inconclusive on a causal role. Genetic studies provide indirect support for a role of mycobacteria in Crohns disease, by identifying susceptibility genes that encode proteins implicated in innate immunity to intracellular bacteria. Clinical trial data support at least a short-term benefit for antimycobacterial therapy in Crohns disease, but the microbial specificity of this response is presently unknown. Summary There appears to be a strong association between M. avium paratuberculosis and Crohns disease, but the causality of this association is unknown. Consequently, the therapeutic implications of this association require further study. A number of critical questions about the biology of M. avium paratuberculosis remain unanswered. Data from studies of this organism, and its interaction with the immune system, can help address proposed reasons for or against a role of M. avium paratuberculosis in the etiology of Crohns disease.

Fine-structure molecular epidemiological analysis of Staphylococcus aureus recovered from cows.

Sixty-three Staphylococcus aureus isolates recovered from bovine sources in the USA and the Republic of Ireland were characterized by multilocus enzyme electrophoresis (MLEE), ribotyping, and random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) typing at two separate laboratories. The S. aureus isolates were assigned by MLEE to 10 electrophoretic types (ETs) (Index of Discrimination, D = 0.779). In contrast, the same isolates were assigned to 13 ribotypes (D = 0.888), and to 12 RAPD types (D = 0.898). A common clone, ET3, of worldwide distribution, was represented by six distinct combinations of ribotypes and RAPD types. S. aureus clones recovered from cows in Ireland were also associated with mastitis in dairy cows in the USA. These findings are consistent with the hypothesis that only a few specialized clones of S. aureus are responsible for the majority of cases of bovine mastitis, and that these clones have a broad geographic distribution.