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InternalMed_Harrison. diagnostic assays and in monitoring test sensitivity was recently highlighted by the emergence in Sweden of a new variant of C. trachomatis containing a deletion that includes the gene targeted by a set of commercial NAATs. By evading detection through this deletion (which would have prompted the initiation of treatment), this strain came to be highly prevalent in some areas of Sweden. While nucleic acid–based tests remain the diagnostic approach of choice for fastidious bacteria, this example serves as a reminder of the need for careful development and ongoing monitoring of molecular diagnostics.


InternalMed_Harrison

InternalMed_Harrison. In contrast, for typical bacterial pathogens for which culture methods are well established, growth-based assays followed by biochemical tests still dominate in the clinical laboratory. Informed by decades of clinical microbiology, these tests have served clinicians well, yet the limitations of growth-based tests—in particular, the delays associated with waiting for growth—have left open opportunities for improvements. Molecular diagnostics, greatly informed by the vast quantity of microbial genome sequences generated in recent years, offers a way forward. First, sequencing studies may identify key genes (or noncoding nucleic acids) that can be developed into targets for clinical assays using PCR or hybridization platforms. Second, sequencing itself may eventually become inexpensive and rapid enough to be performed routinely on clinical specimens, with consequent unbiased detection of pathogens.


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InternalMed_Harrison. In order to adapt nucleic acid detection to diagnostic tests and thus to identify pathogens on a wide scale, sequences must be identified that are conserved enough within a species to identify the diversity of strains that may be encountered in various clinical settings, yet divergent enough to distinguish one species from another. Until recently, this problem has been solved for bacteria by targeting the element of a bacterial genome that is most highly conserved within a species: the 16S ribosomal RNA (rRNA) subunit. At present, 16S PCR amplification from 772 tissue specimens can be performed by specialty laboratories, though its sensitivity and clinical utility to date have remained somewhat limited because, for instance, of inhibitory molecules often found in clinical tissue samples that prevent reliable, sensitive PCR amplification. As such barriers are reduced through technological advances and as the causes of culture-negative infection are clarified (perhaps in part through


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InternalMed_Harrison. that prevent reliable, sensitive PCR amplification. As such barriers are reduced through technological advances and as the causes of culture-negative infection are clarified (perhaps in part through sequencing efforts), these tests may become both more accessible and more helpful. With the wealth of sequencing data now available, other regions beyond 16S rRNA can be targeted for bacterial species identification. These other genomic loci can provide additional information about a clinical isolate that is relevant to patient management. For instance, detection of the presence—or potentially even the expression—of toxin genes such as those for C. difficile toxins A and B or Shiga toxin may provide clinicians with additional information that will help distinguish commensals or colonizing bacteria from pathogens and thus aid in prognostication as well as diagnosis. While amplification tests such as PCR exemplify one approach to nucleic acid detection, other approaches exist, including


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InternalMed_Harrison. bacteria from pathogens and thus aid in prognostication as well as diagnosis. While amplification tests such as PCR exemplify one approach to nucleic acid detection, other approaches exist, including detection by hybridization. Although not currently used in the clinical realm, techniques for detection and identification of pathogens by hybridization to microarrays are being developed for other purposes. Of note, these different detection techniques require different degrees of conservation. Highly sensitive amplification methods require a high degree of sequence identity between PCR primer pairs and their short, specific target sequences; even a single base-pair mismatch (particularly near the 3′ end of the primer) may interfere with detection. In contrast, hybridization-based tests are more tolerant of mismatch and thus can be used to detect important regions that may be less precisely conserved within a species, thus potentially allowing detection of clinical isolates from a given


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InternalMed_Harrison. more tolerant of mismatch and thus can be used to detect important regions that may be less precisely conserved within a species, thus potentially allowing detection of clinical isolates from a given species with greater diversity between isolates. Such assays take advantage of the predictable binding interactions of nucleic acids. The applicability of hybridization-based methods toward either DNA or RNA opens up the possibility of expression profiling, which can uncover phenotypic information from nucleic acid content. Both PCR and hybridization methods target specific, known organisms. At the other extreme, as sequencing costs and turnaround times decrease, direct metagenomic sequencing from patient samples is becoming increasingly feasible. This shotgun sequencing approach is unbiased—i.e., is able to detect any microbial sequence, however divergent or unexpected. This new approach brings its own set of challenges, however, including the need to recognize pathogenic sequences


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InternalMed_Harrison. is able to detect any microbial sequence, however divergent or unexpected. This new approach brings its own set of challenges, however, including the need to recognize pathogenic sequences against a background of expected host and commensal sequences and to distinguish true pathogens from either colonizers or laboratory contaminants. In a powerful example of this new frontier of sequencing-based clinical diagnosis, investigators diagnosed neuroleptospirosis in a child with an unexplained encephalitis syndrome by finding sequences corresponding to the Leptospira genus in cerebrospinal fluid from the patient. Rapid (<48-h) sequencing and analysis informed the patient’s care in real time, leading to life-saving targeted antibiotic therapy for an unexpected diagnosis that was impossible to make through standard laboratory testing. The diagnosis was retrospectively confirmed through both convalescent serologies and PCR using primers designed on the basis of sequencing data.


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InternalMed_Harrison. In addition to clinical diagnostic applications, novel genomic technologies, including whole-genome sequencing, are being applied to clinical research specimens with a goal of identifying new pathogens in a variety of circumstances. The tremendous sensitivity and unbiased nature of sequencing is also ideal in searching clinical samples for unknown or unsuspected pathogens.


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InternalMed_Harrison. Causal inference in infectious diseases has progressed since the time of Koch, whose historical postulates provided a rigorous framework for attributing a disease to a microorganism. According to an updated version of Koch’s postulates, an organism, whether it can be cultured or not, should induce disease upon introduction into a healthy host if it is to be implicated as a causative pathogen. Current sequencing technologies are ideal for advancing this modern version of Koch’s postulates because they can identify candidate causal pathogens with unprecedented sensitivity and in an unbiased way, unencumbered by limitations such as culturability. Yet, as direct sequencing on primary patient samples greatly expands our ability to recognize associations between microbes and disease states, critical thinking and experimentation will continue to be vital to establishing causality.


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InternalMed_Harrison. Virus discovery in particular has been greatly facilitated by new nucleic acid technology. These frontiers were first notably explored with high-density microarrays containing spatially arrayed sequences from a phylogenetically diverse collection of viruses. Although biased toward those with homology to known viruses, novel viruses in clinical samples were successfully identified on the basis of their ability to hybridize to these prespecified sequences. This methodology famously contributed to identification of the coronavirus causing severe acute respiratory syndrome (SARS). Once discovered, this SARS corona-virus was rapidly sequenced: the full genome was assembled in April 2003, less than 6 months after recognition of the first case. This accomplishment illustrated the advancing power and speed of new diagnostic technologies.


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InternalMed_Harrison. With the advent of next-generation sequencing, unbiased pathogen discovery is now possible through a process known as metagenomic assembly (Fig. 146-3). Sequences of random nucleotide fragments can be generated from clinical specimens with no a priori knowledge of pathogen identity through a process called shotgun sequencing. This collection of sequences can then be computationally aligned to host (i.e., human) sequences, with aligned sequences removed and remaining sequences compared with other known genomes to detect the presence of known microorganisms. Sequence fragments that remain unaligned suggest the presence of an additional organism that cannot be matched to a known, characterized genome; these reads can be assembled into contiguous nucleic acid stretches that can be compared to known sequences to construct the genome of a potentially novel organism. Assembled genomes (or parts of genomes) can then be compared to known genomes to infer the phylogeny of new organisms and


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InternalMed_Harrison. to known sequences to construct the genome of a potentially novel organism. Assembled genomes (or parts of genomes) can then be compared to known genomes to infer the phylogeny of new organisms and identify related classes or traits. Thus, not only can this process identify unanticipated pathogens; it can even identify undiscovered organisms. Some early applications of sequencing on clinical samples have centered around the discovery of novel viruses, including such emerging pathogens as West Nile virus, SARS coronavirus, and the Middle East respiratory syndrome coronavirus (MERS-CoV) that has caused severe respiratory illnesses in healthy adults, as well as viral causes of myriad other conditions, from tropical hemorrhagic fevers to diarrhea in newborns.


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InternalMed_Harrison. More recently, metagenomic assembly has been successfully extended to bacterial pathogen discovery. Investigators identified a new bacterial species associated with “cord colitis”—a rare antibiotic-responsive, culture-negative colitis in recipients of umbilical cord-blood stem cells—by sequencing colon biopsy samples from affected patients and matched controls. A single dominant species emerged from metagenomic assembly in samples from patients that was absent from control samples. The presence of this species was confirmed by PCR and fluorescence in situ hybridization on primary tissue samples. On the basis of its similarity to other known species, the organism was named Bradyrhizobium enterica, a novel species from a genus that has proved difficult to culture and thus would have been hard to identify by other means. Correlation versus causation remains an open question; therefore, further efforts will be required to make such links.


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InternalMed_Harrison. As metagenomic sequencing and assembly techniques become more robust, this technology holds great promise for identifying microorganisms that are associated with clinical conditions of unknown etiology. Conventional methods already have unexpectedly linked numerous conditions with specific agents of infection—e.g., cervical and oropharyngeal cancers with human papillomavirus, Kaposi’s sarcoma with human herpesvirus 8, and certain lymphomas with Epstein-Barr virus. Sequencing techniques offer unprecedented sensitivity and specificity for identifying foreign nucleic acid sequences that may suggest other conditions—from malignancies to inflammatory conditions to unexplained fevers or other clinical syndromes—associated with organisms ...CCTAAGGGCTCCAGA + CTCCAGAGTTCAGTC... taxonomic assignment of reads ...CCTAAGGGCTCCAGAGTTCAGTC... host known genomes comparison to


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InternalMed_Harrison. host known genomes comparison to FIGURE 146-3 Workflow of metagenomic assembly for pathogen discovery. DNA is isolated from a specimen of interest (e.g., tissue, body fluid) containing a mixture of host DNA and nucleic acids from coexisting microbes, either commensal or pathogenic. All DNA (and RNA if a reverse transcription step is added) is then sequenced, yielding a mixture of DNA sequence fragments (“reads”) from organisms present. These reads are then aligned to existing reference genomes for the host or any known microbes, leaving reads that do not align (“map”) to any known sequence. These unmapped reads are then computationally assembled de novo into the largest contiguous stretches of DNA possible (“contigs”), representing fragments of previously unsequenced genomes. These genome fragments (contigs) are then mapped onto a phylogenetic tree based on their sequence. Some may represent known but as-yet-unsequenced organisms, while others will represent novel species.


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InternalMed_Harrison. (Figure prepared with valuable input from Dr. Ami S. Bhatt, personal communication.) from viruses to bacteria to parasites. As sequencing-based discovery expands, microbes may be found to be associated with conditions not classically thought of as infectious. Studies of bowel flora in laboratory animals and even humans are already beginning to suggest correlations between microbe composition and various aspects of metabolic and cardiovascular health. Improved methods for pathogen detection will continue to uncover unexpected correlations between microbes and disease states, but the mere presence of a microbe does not establish causality. Fortunately, once the relatively laborious and computationally intensive metagenomic sequencing and assembly efforts have identified a pathogen, further detection can easily be undertaken with targeted methods such as PCR or hybridization, which are much more straightforward and scalable. This capacity should facilitate the additional careful


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InternalMed_Harrison. further detection can easily be undertaken with targeted methods such as PCR or hybridization, which are much more straightforward and scalable. This capacity should facilitate the additional careful investigation that will be required to progress beyond correlation and to draw causal inference.


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InternalMed_Harrison. At present, antibiotic resistance in bacteria and fungi is determined by isolating a single colony from a cultured clinical specimen and testing its growth in the presence of a drug. The requirement for multiple growth steps in these conventional assays has several consequences. First, only culturable pathogens can be readily processed. Second, this process requires considerable infrastructure to support the sterile environment required for culture-based testing of diverse organisms. Finally, and perhaps most significantly, even the fastest-growing organisms require 1–2 days of processing for identification and 2–3 days for determination of susceptibilities. Slower-growing organisms take even longer: for instance, weeks must pass before drug-resistant


InternalMed_Harrison

InternalMed_Harrison. M. tuberculosis can be identified by growth phenotype. Given the clinical imperative in serious illness to begin effective therapy early, this inherent delay in susceptibility determination has obvious implications for empirical antibiotic use: broad-spectrum antibiotics often must be chosen up front in situations where it is later shown that preferred narrower-spectrum drugs would have been effective or even that no antibiotics were appropriate (i.e., in viral infections). With this strategy, the empirical choice can be incorrect, often with devastating consequences. Real-time identification of the infecting organism and information on its susceptibility profile would guide initial therapy and support judicious antibiotic use, ideally improving patient outcomes while aiding in the ever-escalating struggle with antibiotic resistance by reserving the use of broad-spectrum agents for cases in which they are truly needed.


InternalMed_Harrison

InternalMed_Harrison. Molecular diagnostics and sequencing offer a way to accelerate detection of a pathogen’s antibiotic susceptibility profile. If a genotype that confers resistance can be identified, this genotype can be targeted for molecular detection. In infectious disease, this approach has most convincingly come to fruition for HIV (Fig. 146-2A). (In a conceptually parallel application of genomic analysis, molecular detection of certain resistance determinants in cancers is beginning to inform chemotherapeutic selection.) Extensive sequencing of HIV strains and correlations drawn between viral genotypes and phenotypic resistance have delineated the majority of mutations in key HIV genes, such as reverse transcriptase, protease, and integrase, that confer resistance to the antiretroviral agents that target these proteins. For instance, the singleamino-acid substitution K103N in the HIV reverse transcriptase gene predicts resistance to the first-line nonnucleoside reverse transcriptase inhibitor


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InternalMed_Harrison. target these proteins. For instance, the singleamino-acid substitution K103N in the HIV reverse transcriptase gene predicts resistance to the first-line nonnucleoside reverse transcriptase inhibitor efavirenz, and its detection thus informs a clinician to choose a different agent. The effects of these common mutations on HIV susceptibility to various drugs—as well as on viral fitness—are curated in publically available databases. Thus, genotypes are now routinely used to predict drug resistance in HIV, as phenotypic resistance assays are far more cumbersome than targeted sequencing. Indeed, current recommendations in the United States are to sequence virus from a patient’s blood before initiating antiretroviral therapy, which is then tailored to the predicted resistance phenotype. As new targeted therapies are introduced, this targeted sequencing–based approach to drug resistance will likely prove important in other viral infections (e.g., hepatitis C).


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InternalMed_Harrison. For several reasons, the challenge of predicting antibiotic susceptibility from genotype has not yet been met in bacteria to the same degree as in HIV. In general, bacteria have evolved diverse resistance mechanisms to most antibiotics; thus, the task cannot be reduced to probing for a single genetic lesion, target, or mechanism. For instance, at least five distinct modes of resistance to fluoroquinolones are known: reduced import, increased efflux, mutated target sites, drug modification, and shielding of the target sites by expression of another protein. Further, we lack a comprehensive compendium of genetic elements conferring resistance, and new mechanisms and genes emerge regularly in the face of antibiotic deployment. As bacteria have far more complex genomes than viruses, with thousands of genes on their chromosomes and the capacity for acquiring many more through 774 horizontal gene transfer of plasmids and mobile genetic elements within and even between species, the task of


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InternalMed_Harrison. of genes on their chromosomes and the capacity for acquiring many more through 774 horizontal gene transfer of plasmids and mobile genetic elements within and even between species, the task of not only defining all current but also predicting all future mechanisms at a genetic level is daunting or perhaps impossible. Despite these challenges, in a few select cases where the genotypic basis for resistance has been well defined, genotypic assays for antibiotic resistance are already being introduced into clinical practice. One important example is the detection of methicillin-resistant Staphylococcus aureus (MRSA). S. aureus is one of the most common and serious bacterial pathogens of humans, particularly in health care settings. Resistance to methicillin, the most effective antistaphylococcal antibiotic, has become very common even in community-acquired strains. The alternative to methicillin, vancomycin, is effective against MRSA but measurably inferior to methicillin against


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InternalMed_Harrison. antibiotic, has become very common even in community-acquired strains. The alternative to methicillin, vancomycin, is effective against MRSA but measurably inferior to methicillin against methicillinsusceptible S. aureus (MSSA). Analysis of clinical MRSA isolates has demonstrated that the molecular basis for resistance to methicillin in essentially all cases stems from the expression of an alternative penicillin-binding protein (PBP2A) encoded by the gene mecA, which is found within a transferable genetic element called mec. This mobile cassette has spread rapidly through the S. aureus population via horizontal gene transfer and selection from widespread antibiotic use. Because resistance is essentially always due to the presence of the mec cassette, MRSA is amenable to molecular detection. In recent years, a PCR test for the presence of the mec cassette, which saves hours to days compared with standard culture-based methods, has been approved by the U.S. Food and Drug Administration.


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InternalMed_Harrison. In recent years, a PCR test for the presence of the mec cassette, which saves hours to days compared with standard culture-based methods, has been approved by the U.S. Food and Drug Administration. Additional molecular diagnostics are being implemented in the evaluation of bacterial antibiotic resistance. Vancomycin-resistant enterococci (VRE) harbor one of a limited number of van genes responsible for resistance to this important antibiotic by altering the mechanism for cell wall cross-linking that vancomycin inhibits. Detection of one of these genes by PCR indicates resistance. Identification of two carbapenemase-encoding plasmids—NDM-1 and KPC—responsible for a significant fraction of carbapenem resistance (though not for all such resistance) has led to the development of a multiplexed PCR assay to detect these important resistance elements. Because carbapenems are broad-spectrum antibiotics frequently reserved for multidrugresistant bacteria (particularly enteric gram-negative


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InternalMed_Harrison. PCR assay to detect these important resistance elements. Because carbapenems are broad-spectrum antibiotics frequently reserved for multidrugresistant bacteria (particularly enteric gram-negative bacilli) and are often used as antibiotics of last resort, the initial appearance of resistance and the subsequent increase in its prevalence have caused considerable concern. Therefore, even though other mechanisms for carbapenem resistance exist, a rapid PCR test for the two plasmids encoding these two carbapenemases has been developed to aid in both diagnostic and infection control efforts. Efforts are under way to extend this multiplexed PCR assay to other plasmid-borne carbapenemases and thus to make it more comprehensive. The power and application of molecular genetic tests are not limited to high-income settings. With the increasing burden of drug-resistant tuberculosis in the developing world, a molecular diagnostic test has now been developed to detect rifampin-resistant


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InternalMed_Harrison. not limited to high-income settings. With the increasing burden of drug-resistant tuberculosis in the developing world, a molecular diagnostic test has now been developed to detect rifampin-resistant tuberculosis. The genetic basis for rifampin resistance has been well defined by targeted sequencing: characteristic mutations in the molecular target of rifampin, RNA polymerase, account for the vast majority of rifampinresistant strains of M. tuberculosis. A PCR assay that can detect both the M. tuberculosis organism and a rifampin-resistant allele of RNA polymerase from clinical samples has recently been approved. Since rifampin resistance frequently accompanies resistance to other antibiotics, this test can suggest the possible presence of multidrug-resistant


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InternalMed_Harrison. M. tuberculosis within hours instead of weeks. Despite differences in relative genome complexity, HIV genotypic screening for antiretroviral resistance offers one framework for broadening efforts at genotypic assays for nonviral antibiotic resistance. As whole-genome pathogen sequencing has become increasingly feasible and inexpensive (Fig. 146-1), significant efforts have been launched to sequence hundreds to thousands of antibiotic-sensitive and -resistant isolates of a given pathogen in order to more comprehensively define resistance-conferring genetic elements. In parallel with advancing sequencing technologies, progress in computational techniques, bioinformatics and statistics, and data storage as well as experimental confirmatory testing of hypotheses will be needed to move toward the ambitious goal of a comprehensive compendium of antibiotic resistance determinants. Open sharing and careful curation of new sequence information will be of paramount importance.


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InternalMed_Harrison. Yet no matter how thorough and carefully curated such a genotype-phenotype database is, history suggests that comprehensively cataloguing resistance in nonviral pathogens, with new mechanisms continuously emerging, will be challenging at best. Even identifying and itemizing current resistance mutations is a daunting prospect: nonviral genomes are much larger than viral ones, and their abundance and diversity are such that hundreds to thousands of genetic differences often exist between clinical isolates, of which perhaps only one may cause resistance. For example, increasing resistance to artemisinin, one of the most effective new agents for malaria, has prompted recent large-scale efforts to identify the basis for resistance. While such studies have identified promising leads, no clear mechanism has emerged; in fact, a single genetic lesion alone may not fully account for resistance. Especially with multiple possible resistance mechanisms for a given antibiotic as well as ongoing


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InternalMed_Harrison. mechanism has emerged; in fact, a single genetic lesion alone may not fully account for resistance. Especially with multiple possible resistance mechanisms for a given antibiotic as well as ongoing evolutionary pressure resulting in the development and acquisition of new modes of resistance, a genotypic approach to diagnosing antibiotic resistance is likely to be imperfect.


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InternalMed_Harrison. We have already observed the accumulation of new or unanticipated modes of resistance from ongoing evolutionary pressure caused by the widespread clinical use of antibiotics. Even with MRSA, perhaps the best-studied case of antibiotic resistance and a model of relative simplicity with a single known monogenic resistance determinant (mecA), a genotype-based approach to resistance detection proved flawed. One limitation was a recall of the initial commercial genotypic resistance assay that was deployed for the identification of MRSA. A clinical isolate of S. aureus emerged in Belgium that expressed a variant of the mec cassette not detected by the assay’s PCR primers. New primers were added to detect this new variant, and the assay was re-approved for use. More recently, an even more divergent but functionally analogous gene called mecC, which confers methicillin resistance but evades PCR detection by this assay, was found. This series of events exemplifies the need for ongoing


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InternalMed_Harrison. divergent but functionally analogous gene called mecC, which confers methicillin resistance but evades PCR detection by this assay, was found. This series of events exemplifies the need for ongoing monitoring of any genotypic resistance assay. A second limitation is that a contradiction can occur between genotypic and phenotypic evidence for resistance. Up to 5% of MSSA strains carry a copy of the mecA gene that is either nonfunctional or not expressed. Thus, the erroneous identification of these strains as MRSA by genotypic detection would lead to administration of the inferior antibiotic vancomycin rather than the preferred β-lactam therapy.


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InternalMed_Harrison. These examples illustrate one of the prime challenges of moving beyond growth-based assays: genotype is merely a proxy for the resistance phenotype that directly informs patient care. One alternative approach currently under development attempts to circumvent the limitations of genotypic resistance testing by returning to a phenotypic approach, albeit one informed by genomic methods: transcriptional profiles serve as a rapid phenotypic signature for antibiotic response. Conceptually, since dying cells are transcriptionally distinct from cells fated to survive, susceptible bacteria enact different transcriptional profiles after antibiotic exposure that are different from the profiles of resistant strains, independent of the mechanism of resistance. These differences can be measured and, since transcription is one of the most rapid responses to cell stress (minutes to hours), can be used to determine whether cells are resistant or susceptible much more rapidly than is possible if one


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InternalMed_Harrison. since transcription is one of the most rapid responses to cell stress (minutes to hours), can be used to determine whether cells are resistant or susceptible much more rapidly than is possible if one waits for growth in the presence of antibiotics (days). Like DNA, RNA can be readily detected through predictable rules governing base pairing via either amplification or hybridization-based methods. Changes in a carefully selected set of transcripts form an expression signature that can represent the total cellular response to antibiotic without requiring full characterization of the entire transcriptome. Preliminary proof-of-concept studies suggest that this approach may identify antibiotic susceptibility on the basis of transcriptional phenotype much more quickly than is possible with growth-based assays.


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InternalMed_Harrison. Because of its sensitivity in detecting even very rare nucleic acid fragments, sequencing is now permitting studies of unprecedented depth into complex populations of cells and tissues. The strength of this depth and sensitivity applies not only to the detection of rare, novel pathogens in a sea of host signal but also to the identification of heterogeneous pathogen subpopulations in a single host that may differ, for example, in drug resistance profiles or pathogenesis determinants. Future studies will be needed to elucidate the clinical significance of these variable subpopulations, even as deep sequencing is now providing unprecedented levels of detail about majority and minority members of this population.


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InternalMed_Harrison. While pathogen-based diagnostics continue to be the mainstay for diagnosing infection, serologic testing has long been the basis of a strategy to diagnose infection by measuring host responses. Here, too, the application of genomics is now being explored to improve upon this approach, given the previously described limitations of serologic testing. Rather than using antibody responses as a retrospective biomarker for infection, recent efforts have focused on transcriptomic analysis of the host response as a new direction with diagnostic implications for human disease. For instance, while pathogen-based diagnostic tests to distinguish active from latent tuberculosis infection have proved elusive, recent work shows that the transcriptional profile of circulating white blood cells exhibits a differential pattern of expression of nearly 400 transcripts that distinguish active from latent tuberculosis; this expression pattern is driven in part by changes in interferon-inducible genes in


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InternalMed_Harrison. a differential pattern of expression of nearly 400 transcripts that distinguish active from latent tuberculosis; this expression pattern is driven in part by changes in interferon-inducible genes in the myeloid lineage. In a validation cohort, this transcriptional signature was able to distinguish patients with active versus latent disease, to distinguish tuberculosis infection from other pulmonary inflammatory states or infections, and to track responses to treatment in as little as 2 weeks, with normalization of expression toward that of patients without active disease over 6 months of effective therapy. Such a test could play an important role not only in the management of patients but also as a marker of efficacy in clinical trials of new therapeutic agents. Similarly, other investigators have been trying to identify host transcriptional signatures in circulating blood cells that are distinct in influenza A infection from those in upper respiratory infections caused by certain


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InternalMed_Harrison. have been trying to identify host transcriptional signatures in circulating blood cells that are distinct in influenza A infection from those in upper respiratory infections caused by certain other viruses or bacteria. These signatures also varied with phase of infection and showed promise in distinguishing exposed subjects who will become symptomatic from those who will not. These results suggest that profiling of host transcriptional dynamics could augment the information obtained from studies of pathogens, both enhancing diagnosis and monitoring the progression of illness and the response to therapy.


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InternalMed_Harrison. In this era of genome-wide association studies and attempts to move toward personalized medicine, genomic approaches are also being applied to the identification of host genetic loci and factors that contribute to infection susceptibility. Such loci will have undergone strong selection among populations in which the disease is endemic. By identifying the beneficial genetic alleles among individuals who survive in such settings, markers for susceptibility or resistance are being discovered; these markers can be translated into diagnostic tests to identify susceptible individuals in order to implement preventive or prophylactic interventions. Further, such studies may offer mechanistic insight into the pathogenesis of infection and inform new methods of therapeutic intervention. Such beneficial genetic associations were recognized long before the advent of genomics, as in the protective effects of the negative Duffy blood group or heterozygous hemoglobin abnormalities against Plasmodium


InternalMed_Harrison

InternalMed_Harrison. genetic associations were recognized long before the advent of genomics, as in the protective effects of the negative Duffy blood group or heterozygous hemoglobin abnormalities against Plasmodium infection. Genomic methods enable more systematic and widespread investigations of the host to identify not only people with altered susceptibility to numerous diseases (e.g., HIV infection, tuberculosis, and cholera) but also host factors that contribute to and thus might predict the severity of disease.


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InternalMed_Harrison. Genomics has the potential to impact infectious disease therapeutics in two ways. By transforming the speed of diagnostic information acquisition or the type of diagnostic information that can be attained, it can influence therapeutic decision-making. Alternatively, by opening up new avenues to understanding pathogenesis, providing new ways to disrupt infection, and delineating new approaches to antibiotic 775 discovery, it can facilitate the development of new therapeutic agents.


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InternalMed_Harrison. Efforts at antibiotic discovery are declining, with few new agents in the pipeline and even fewer entering the market. This phenomenon is due in part to the lack of economic incentives for the private sector; however, it is also attributable in part to the enormous challenges involved in the discovery and development of antibiotics. For obvious market-related reasons, nearly all efforts have focused on broad-spectrum antibiotics; the development of a chemical entity that works across an extremely diverse set of organisms (i.e., more divergent from each other than a human is from an amoeba) is far more challenging than the development of an agent that is designed to target a single bacterial species. Nevertheless, the concept of narrow-spectrum antibiotics has heretofore been rejected because of the lack of early diagnostic information that would guide the selection of such agents. Thus, rapid diagnostics providing antibiotic susceptibility information that can guide antibiotic


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InternalMed_Harrison. because of the lack of early diagnostic information that would guide the selection of such agents. Thus, rapid diagnostics providing antibiotic susceptibility information that can guide antibiotic selection in real time have the potential to alter and simplify antibiotic strategies by allowing a paradigm shift away from broad-spectrum drugs and toward narrow-spectrum agents. Such a paradigm shift clearly would have additional implications for the current escalation of antibiotic resistance.


InternalMed_Harrison

InternalMed_Harrison. In yet another diagnostic paradigm with the potential to impact therapeutic interventions, genomics is opening new avenues to a better understanding not only of different host susceptibilities to infection but also of different host responses to therapy. In a sense, the promise of “personalized medicine” has been a tantalizing holy grail. Some signs now point to the realization of this goal. For example, the role of glucocorticoids in tuberculous meningitis has been long debated. Recently, polymorphisms in the human genetic locus LTA4H, which encodes a leukotriene-modifying enzyme, were found to modulate the inflammatory response to tuberculosis. Patients with tuberculous meningitis who were homozygous for the proinflammatory LTA4H allele were most helped by adjunctive glucocorticoid treatment, while those who were homozygous for the anti-inflammatory allele were negatively affected by steroid treatment. This anti-inflammatory adjunct has become the standard of care in tuberculous


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InternalMed_Harrison. while those who were homozygous for the anti-inflammatory allele were negatively affected by steroid treatment. This anti-inflammatory adjunct has become the standard of care in tuberculous meningitis, but this study suggests that perhaps only a subset of patients benefit and further suggests a genetic means of prospectively identifying this subset. Thus, genomic diagnostic tests may eventually inform diagnosis, prognosis, and treatment decisions by revealing the pathogenic potential of the microbe and detecting host responses to both infection and therapy.


InternalMed_Harrison

InternalMed_Harrison. Genomic technologies are already dramatically changing research on host–pathogen interactions, with a goal of increasingly influencing the process of therapeutic discovery and development. Sequencing offers several possible avenues into antimicrobial therapeutic discovery. First, genomic-scale molecular methods have paved the way for comprehensive identification of all essential genes encoded within a pathogen’s genome, with consequent systematic identification of all possible vulnerabilities within a pathogen that could be targeted therapeutically. Second, transcriptional profiling can offer insights into mechanisms of action of new candidate drugs that emerge from screens. For instance, the transcriptional signature of cell wall disruptors (e.g., β-lactams) is distinct from that of DNA-damaging agents (e.g., fluoroquinolones) or protein synthesis inhibitors (e.g., aminoglycosides). Thus, transcriptional analysis of a pathogen’s response to a new antibiotic can either suggest a


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InternalMed_Harrison. DNA-damaging agents (e.g., fluoroquinolones) or protein synthesis inhibitors (e.g., aminoglycosides). Thus, transcriptional analysis of a pathogen’s response to a new antibiotic can either suggest a mechanism of action or flag compounds for prioritization because of a potentially novel activity. In an alternative genomic strategy for determining mechanisms of action, an RNA interference approach followed by targeted sequencing has been used to identify genes required for antitrypanosomal drug efficacy. This approach provided new insights into the mechanism of action of drugs that have been in use for decades for human African trypanosomiasis. Third, sequencing can readily identify the most conserved regions of a pathogen’s genomes and corresponding gene products; this information is invaluable in narrowing antigen candidates for vaccine development. These surface 776 proteins can be expressed recombinantly and tested for the ability to elicit a serologic response and protective


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InternalMed_Harrison. invaluable in narrowing antigen candidates for vaccine development. These surface 776 proteins can be expressed recombinantly and tested for the ability to elicit a serologic response and protective immunity. This process, termed reverse vaccinology, has proved particularly useful for pathogens that are difficult to culture or poorly immunogenic, as was the case with the development of a vaccine for Neisseria meningitidis serogroup B. Large-scale gene content analysis from sequencing or expression profiling enables new research directions that provide novel insights into the interplay of pathogen and host during infection or colonization. One important goal of such research is to suggest new therapeutic approaches to disrupt this interaction in favor of the host. Indeed, one of the most immediate applications of next-generation sequencing technology has come from simply characterizing human pathogens and related commensal or environmental strains and then finding genomic correlates


InternalMed_Harrison

InternalMed_Harrison. immediate applications of next-generation sequencing technology has come from simply characterizing human pathogens and related commensal or environmental strains and then finding genomic correlates for pathogenicity. For instance, as Escherichia coli varies from a simple nonpathogenic, lab-adapted strain (K-12) to a Shiga toxin–producing enterohemorrhagic gastrointestinal pathogen (O157:H7), it displays up to a 25% difference in gene content, even though its phylogenetic classification stays the same. Although this is an extreme example, it is not an isolated case. Some isolates of Enterococcus—notorious for its increasing incidence of resistance to common antibiotics such as ampicillin, vancomycin, and aminoglycosides—also contain recently acquired genetic material comprising up to 25% of the genome on mobile genetic elements. This fact suggests that horizontal gene transfer may play an important role in the organism’s adaptation as a nosocomial pathogen. On closer study, this


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InternalMed_Harrison. 25% of the genome on mobile genetic elements. This fact suggests that horizontal gene transfer may play an important role in the organism’s adaptation as a nosocomial pathogen. On closer study, this genome expansion has been demonstrated to be associated with loss of regulatory elements called CRISPRs (clustered, regularly interspaced short palindromic repeats). Loss of CRISPR elements, which protect the bacterial genome from invasion by certain foreign genetic materials, may thus facilitate the acquisition of antibiotic resistance–conferring genetic elements. While loss of this regulation appears to impose a competitive disadvantage in antibiotic-free environments, these drug-resistant strains thrive in the presence of even some of the most useful antienterococcal therapies. In addition to insights gained from genome sequencing, extension of unbiased whole-transcriptome sequencing (RNA-Seq) efforts to bacteria is beginning to identify unexpected regulatory, noncoding RNAs in many


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InternalMed_Harrison. to insights gained from genome sequencing, extension of unbiased whole-transcriptome sequencing (RNA-Seq) efforts to bacteria is beginning to identify unexpected regulatory, noncoding RNAs in many diverse species. While the functional implications of these new transcripts are as yet largely unknown, the presence of such features—conserved across many bacterial species— implies evolutionary importance and suggests areas for future study and possible new therapeutic avenues. Thus, genomic studies are already beginning to transform our understanding of infection, offering evidence of virulence factors or toxins and providing insight into ongoing evolution of pathogenicity and drug resistance. One goal of such studies is to identify therapeutic agents that can disrupt the pathogenic process; there is currently much interest in the theoretical concept of antivirulence drugs that inhibit virulence factors rather than killing the pathogen outright as a means to intervene in infection.


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InternalMed_Harrison. there is currently much interest in the theoretical concept of antivirulence drugs that inhibit virulence factors rather than killing the pathogen outright as a means to intervene in infection. Further, as sequencing becomes increasingly accessible and efficient, large-scale studies with unprecedented statistical power to associate clinical outcomes with pathogen and host genotypes and thus to further reveal vulnerabilities in the infection process that can be targeted for disruption are being initiated. Although this is just the beginning, such studies point to a tantalizing future in which the clinician is armed with genomic predictors of infection outcome and therapeutic response to guide clinical decision-making.


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InternalMed_Harrison. Epidemiologic studies of infectious diseases have several main goals: to identify and characterize outbreaks, to describe the pattern and dynamics of an infectious disease as it spreads through populations, and to identify interventions that can limit or reduce the burden of disease. One classic, paradigmatic example is John Snow’s elucidation of the origin of the 1854 London cholera outbreak. Snow used careful geographic mapping of cases to determine that the likely source of the outbreak was contaminated water from the Broad Street pump, and, by removing the pump handle, he aborted the outbreak. Whereas that intervention was undertaken without knowledge of the causative agent of cholera, advances in microbiology and genomics have expanded the purview of epidemiology, which now considers not just the disease but also the pathogen, its virulence factors, and the complex relationships between microbial and host populations.


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InternalMed_Harrison. Through the use of novel genomic tools such as high-throughput sequencing, the diversity of a microbial population can now be rapidly described with unprecedented resolution, with discrimination between isolates that have single-nucleotide differences across the entire genome and advancement beyond prior approaches that relied on phenotypes (such as antibiotic resistance testing) or genetic markers (such as multilocus sequence typing). The development of statistical methods grounded in molecular genetics and evolutionary theory has established analytical approaches that translate descriptions of microbial population diversity and structure into insights into the origin and history of pathogen spread. By linking phylogenetic reconstruction with epidemiologic and demographic data, genomic epidemiology provides the opportunity to track transmission from person to person, to infer transmission patterns of both pathogens and sequence elements that confer phenotypes of interest, and to


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InternalMed_Harrison. epidemiology provides the opportunity to track transmission from person to person, to infer transmission patterns of both pathogens and sequence elements that confer phenotypes of interest, and to estimate the transmission dynamics of outbreaks.


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InternalMed_Harrison. The use of comparisons of whole-genome sequencing to infer person-to-person transmission and identify point-source outbreaks of pathogens has proved useful in hospital infection control settings. As reported in a seminal paper in 2010, a study of MRSA in a Thai hospital demonstrated that whole-genome sequencing can be used to infer transmission of a pathogen from patient to patient within a hospital setting through integration of the analysis of accumulation of mutations over time with dates and hospital locations of the infections. Since that time, multiple instances of the use of whole-genome sequencing to define and motivate interventions aimed at interrupting transmission chains have been reported. In another MRSA outbreak in a special-care baby unit in Cambridge, United Kingdom, whole-genome sequencing extended the traditional infection control analysis, which relies on typing organisms by their antibiotic susceptibilities, to sequencing of isolates from clinical samples. This


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InternalMed_Harrison. sequencing extended the traditional infection control analysis, which relies on typing organisms by their antibiotic susceptibilities, to sequencing of isolates from clinical samples. This approach identified an otherwise unrecognized outbreak of a specific MRSA strain that was occurring against a background of the usual pattern of infections caused by a diverse circulating population of MRSA strains. The analysis showed evidence of transmission among mothers within the special-care baby unit and in the community and demonstrated the key role of MRSA carriage in a single health care provider in the persistence of the outbreak. MRSA decolonization of the health care provider terminated the outbreak. In yet another example, in response to the observation of 18 cases of infection by carbapenemase-producing Klebsiella pneumoniae over 6 months at the National Institutes of Health Clinical Research Center, genome sequencing of the isolates was used to discriminate between the possibilities


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InternalMed_Harrison. Klebsiella pneumoniae over 6 months at the National Institutes of Health Clinical Research Center, genome sequencing of the isolates was used to discriminate between the possibilities that these cases represented multiple, independent introductions into the health care system or a single introduction with subsequent transmission. On the basis of network and phylogenetic analysis of genomic and epidemiologic data, the authors reconstructed the likely relationships among the isolates from patient to patient, demonstrating that the spread of resistant Klebsiella infection was in fact due to nosocomial transmission of a single strain.


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InternalMed_Harrison. Uncovering of unexpected transmission events by genomic epidemiology studies is motivating renewed questioning of pathogen ecology and modes of transmission. For example, the rise in prevalence of infections with nontuberculous mycobacteria, including Mycobacterium abscessus, among patients with cystic fibrosis (CF) has led to speculation about the possible role of patient-to-patient transmission in the CF community; however, conventional typing approaches have lacked the resolution to define population structure accurately, a critical component of inferring transmission. Past infection control guidelines discounted the possibility of acquisition of nontuberculous mycobacteria in health care settings, as no strong evidence for such transmission had been described. In a whole-genome sequencing study of M. abscessus isolates from patients with CF, an analytical approach using genome sequencing, epidemiology, and Bayesian modeling examined the likelihood of transmission between patients


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InternalMed_Harrison. study of M. abscessus isolates from patients with CF, an analytical approach using genome sequencing, epidemiology, and Bayesian modeling examined the likelihood of transmission between patients within a CF center; the authors found nearly identical isolates in a number of patients and observed that these isolates were less diverse than isolates from a single individual. Because no clear epidemiologic link places the infected patients in the same place at the same time, this finding highlights a need to explore preexisting notions of circumstances required for transmission and a reconsideration of M. abscessus infection control guidelines. Similar studies of other pathogens—particularly those that share human, other animal host, and environmental reservoirs—will continue to advance our insight into the relative roles and prominence of sources of infection as well as the modes of spread through populations, thereby establishing evidence-based strategies for prevention and intervention.


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InternalMed_Harrison. As increasing numbers of studies aim to carefully define the origins and spread of infectious agents using the high-resolution lens of whole-genome sequencing, fundamental questions are arising with regard to our understanding of infection in a single individual and the process of a single transmission event. For example, a better understanding of a pathogen population’s diversity within a single infected individual is a critical component in interpreting the relationship among isolates from different patients. While we have traditionally thought of individuals as infected with a single bacterial strain, a recent sequencing study of multiple colonies of S. aureus from a single individual showed a “cloud” of diversity; this finding raises a number of questions that will be important to address as this field develops: What is the clinical significance of this diversity? What are the processes that generate and limit diversity? What amount of diversity is transmitted under different


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InternalMed_Harrison. address as this field develops: What is the clinical significance of this diversity? What are the processes that generate and limit diversity? What amount of diversity is transmitted under different conditions and routes of transmission? How do the answers to these questions vary by infectious organism, type of infection, and host and in response to treatment? More comprehensive descriptions of diversity, population dynamics, transmission bottlenecks, and the forces that shape and influence the growth and spread of microbial populations will be a critically important focus of future investigations.


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InternalMed_Harrison. In addition to reconstructing the transmission chains of local outbreaks, genomics-based epidemiologic methods are providing insight into broad-scale geographic and temporal spread of pathogens. A classic example has been the study of cholera, the dehydrating diarrheal illness caused by infection with Vibrio cholerae. Cholera first spread worldwide from the Indian subcontinent in the 1800s and has since caused seven pandemics; the seventh pandemic has been ongoing since the 1960s. An investigation into the geographic patterns of cholera spread in the seventh pandemic used genome sequences from 777 a global collection of 154 V. cholerae strains representing isolates from 1957–2010. This investigation revealed that the seventh pandemic has comprised at least three overlapping waves spreading out from the Indian subcontinent (Fig. 146-4). Further, analysis of the genome of an isolate of V. cholerae from the 2010 outbreak of cholera in Haiti showed it to be more closely related to


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InternalMed_Harrison. out from the Indian subcontinent (Fig. 146-4). Further, analysis of the genome of an isolate of V. cholerae from the 2010 outbreak of cholera in Haiti showed it to be more closely related to isolates from South Asia than to isolates from neighboring Latin America, a result supporting the hypothesis that the outbreak was derived from V. cholerae introduced into Haiti by human travel (likely from Nepal) rather than by environmental or more geographically proximal sources. A subsequent study that dated the time to the most recent common ancestor of a population of V. cholerae isolates from Haiti provided further support for a single point-source introduction from Nepal.


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InternalMed_Harrison. Increasing numbers of investigations into the spread of many pathogens—thus far including strains of S. aureus, S. pneumoniae, Chlamydia, Salmonella, Shigella, E. coli, C. difficile, West Nile virus, rabies virus, and dengue virus—are contributing to a growing atlas of maps describing routes, patterns, and tempos of microbial diversification and dissemination. Large-scale efforts like the 100K Foodborne Pathogen Genome Project, which aims to sequence the genomes of 100,000 strains of food-borne pathogens collected from sources including food, the environment, and farm animals, are possible because of advances in sequencing technologies. Such studies will yield a vast amount of data that can be used to investigate diversity and microbiologic links within distinct niches and the patterns of spread from one niche to another. The increasingly broad adoption of genome sequencing by health care and public health institutions will ensure that the available catalog of genome sequences and


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InternalMed_Harrison. of spread from one niche to another. The increasingly broad adoption of genome sequencing by health care and public health institutions will ensure that the available catalog of genome sequences and associated epidemiologic data will grow very rapidly. With higher-resolution description of microbial diversity and of the dynamics of that diversity over time and across epidemiologic and demographic boundaries and evolutionary niches, we will gain even greater insights into the relationships of transmission routes and patterns of historic spread.


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InternalMed_Harrison. Defining pathogen transmissibility is a critical step in the development of public health surveillance and intervention strategies as this information can help predict the epidemic potential of an outbreak. Transmissibility can be estimated by a variety of methods, including inference from the growth rate of an epidemic together with the generation time of an infection (the mean interval between infection of an index case and of the people infected by that index case). Genome


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InternalMed_Harrison. FIGURE 146-4 Transmission events inferred from phylogenetic reconstruction of 154 Vibrio cholerae isolates from the seventh cholera pandemic. Date ranges represent estimated time to the most recent common ancestor for strains transmitted from source to destination locations, based on a Bayesian model of the phylogeny. (Reprinted with permission from A Mutreja et al: Evidence for several waves of global transmission in the seventh cholera pandemic. Nature 477:462, 2011.) 778 sequencing and analysis of a well-sampled population provide another method by which to derive similar fundamental epidemiologic parameters. One key measure of transmissibility is the basic reproduction number (R0), defined as the number of secondary infections generated from a single primary infectious case. When the basic reproduction number is greater than 1 (R0 >1), an outbreak has epidemic potential; when it is less than 1 (R0 <1), the outbreak will become extinct. On the basis of sequences from influenza


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InternalMed_Harrison. the basic reproduction number is greater than 1 (R0 >1), an outbreak has epidemic potential; when it is less than 1 (R0 <1), the outbreak will become extinct. On the basis of sequences from influenza samples obtained from infected patients very early in the 2009 H1N1 influenza pandemic, the basic reproduction number was estimated through a population genomic analysis at 1.2; this compared to estimates of 1.4–1.6 based on several epidemiologic analyses. In addition, with the assumption of a molecular clock model, sequences of H1N1 samples together with information about when and where the samples were obtained have been used to estimate the date and location of the pandemic’s origin, providing insight into disease origins and dynamics. Because the magnitude and intensity of the public health response are guided by the predicted size of an outbreak, the ability of genomic methods to elucidate a pathogen’s origin and epidemic potential adds an important dimension to the contributions of


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InternalMed_Harrison. response are guided by the predicted size of an outbreak, the ability of genomic methods to elucidate a pathogen’s origin and epidemic potential adds an important dimension to the contributions of these methods to infectious disease epidemiology.


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InternalMed_Harrison. Beyond describing transmission and dynamics, pathogen genomics can provide insight into the evolution of pathogens and the interaction of selective pressures, the host, and pathogen populations, which can have implications for vaccine or therapeutic development. From a clinical perspective, this process is central to the acquisition of antibiotic resistance, the generation of increasing pathogenicity or new virulence traits, the evasion of host immunity and clearance (leading to chronic infection), and vaccine efficacy.


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InternalMed_Harrison. Microbial genomes evolve through a variety of mechanisms, including mutation, duplication, insertion, deletion, recombination, and horizontal gene transfer. Segmented viruses (e.g., influenza virus) can reassort gene segments within multiply infected cells. The pandemic 2009 H1N1 influenza A virus, for example, appears to have been generated through reassortment of several avian, swine, and human influenza strains. Such potential for the evolution of novel pandemic strains has precipitated concern about the possible evolution to transmissibility of virulent strains that have been associated with high mortality rates but have not yet exhibited efficient human infectivity. Controversial experiments with H5N1 avian influenza virus, for example, have defined five mutations that render the virus transmissible, at least in ferrets—the animal model system for human influenza.


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InternalMed_Harrison. The continual antigenic evolution of seasonal influenza offers an example of how studies of pathogen evolution can impact surveillance and vaccine development. Frequent updates to the annual influenza vaccine are needed to ensure protection against the dominant strains. These updates are based on an ability to anticipate which viral populations from a pool of substantial locally and globally diverse circulating viruses will predominate in the upcoming season. Toward that end, sequencing-based studies of influenza virus dynamics have shed light on the global spread of influenza, providing concrete data on patterns of spread and helping to elucidate the origins, emergence, and circulation of novel strains. Through analysis of more than 1000 influenza A H3N2 virus isolates over the 2002–2007 influenza seasons, Southeast Asia was identified as the usual site from which diversity originates and spreads worldwide. Further studies of global isolate collections have shed further light on the


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InternalMed_Harrison. influenza seasons, Southeast Asia was identified as the usual site from which diversity originates and spreads worldwide. Further studies of global isolate collections have shed further light on the diversity of circulating virus, showing that some strains persist and circulate outside of Asia for multiple seasons.


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InternalMed_Harrison. Not only do genomic epidemiology studies have the potential to help guide vaccine selection and development; they are also helping to track what happens to pathogens circulating in the population in response to vaccination. By describing pathogen evolution under the selective pressure of a vaccinated population, such studies can play a key role in surveillance and identification of virulence determinants and perhaps may even help to predict the future evolution of escape from vaccine protection. The 7-valent pneumococcal conjugate vaccine (PCV-7) targeted the seven serotypes of S. pneumoniae responsible for the majority of cases of invasive disease at the time of its introduction in 2000; since then, PCV-7 has dramatically reduced the incidence of pneumococcal disease and mortality. Population genomic analysis of the sequences of more than 600 Massachusetts pneumococcal isolates from 2001–2007 has shown that preexisting rare nonvaccine serotypes are replacing vaccine serotypes and


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InternalMed_Harrison. genomic analysis of the sequences of more than 600 Massachusetts pneumococcal isolates from 2001–2007 has shown that preexisting rare nonvaccine serotypes are replacing vaccine serotypes and that some strains have persisted despite vaccination by recombining the vaccine-targeted capsule locus with a cassette of capsule genes from non-vaccine-targeted serotypes.


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InternalMed_Harrison. While cutting-edge genomic technologies are largely imple mented in the developed world, their application to infectious diseases offers perhaps the biggest potential impact in less developed regions where the burden of these infections is greatest. This globalization of genomic technology and its extensions has already begun in each of the areas of focus highlighted in this chapter; it has occurred both through the application of advanced technologies to samples collected in the developing world and through the adaptation and importation of technologies directly to the developing world for on-site implementation as they become more globally accessible. Genomic characterization of the pathogens responsible for important global illnesses such as tuberculosis, malaria, trypanosomiasis, and cholera has led to insights in diagnosis, treatment, and infection control. For instance, the nucleic acid–based test developed for rapid diagnosis of M. tuberculosis infection and detection of


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InternalMed_Harrison. and cholera has led to insights in diagnosis, treatment, and infection control. For instance, the nucleic acid–based test developed for rapid diagnosis of M. tuberculosis infection and detection of rifampin resistance is being priced for implementation in field settings in Africa and Asia where tuberculosis is most prevalent. The potential to diagnose multidrug-resistant tuberculosis in hours instead of weeks or months may truly revolutionize treatment and control of this common and devastating illness. High-resolution genomic tracking of the spread of cholera has yielded insights into which public health measures may prove most effective in controlling local epidemics. Overall, sequencing efforts have become exponentially cheaper with each passing year. As these technologies synergize with efforts to globalize information-technology resources, global implementation of genomic methods promises to spread state-of-the-art methods for diagnosis, treatment, and epidemic tracking of


InternalMed_Harrison

InternalMed_Harrison. with efforts to globalize information-technology resources, global implementation of genomic methods promises to spread state-of-the-art methods for diagnosis, treatment, and epidemic tracking of infections to areas that need these capabilities the most.


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InternalMed_Harrison. By illuminating the genetic information that encodes the most fundamental processes of life, genomic technologies are transforming many aspects of medicine. In infectious diseases, methods such as next-generation sequencing and genome-scale expression analysis offer information of unprecedented depth about individual microbes as well as microbial communities. This information is expanding our understanding of the interactions of these microorganisms and communities with one another, with their human hosts, and with the environment. Despite significant progress and the abundant genomic data now available, technological and financial barriers continue to impede the widespread adoption of large-scale pathogen sequencing in clinical, public health, and research settings. As even vaster amounts of data are generated, innovations in storage, development of bioinformatics tools to manipulate the data, standardization of methods, and training of end-users in both the research and clinical


InternalMed_Harrison

InternalMed_Harrison. of data are generated, innovations in storage, development of bioinformatics tools to manipulate the data, standardization of methods, and training of end-users in both the research and clinical realms will be required. The cost-effectiveness and applicability of whole-genome sequencing, particularly in the clinic, remain to be studied, and studies of the impact of genome sequencing on patient outcomes will be needed to clarify the contexts in which these new methodologies can make the greatest contributions to patient well-being. The ongoing efforts to overcome limitations through collaboration, teaching, and reduction of financial obstacles should be applauded and expanded. With advances in genomic technologies and computational analysis, our ability to detect, characterize, treat, monitor, prevent, and control infections has progressed rapidly in recent years and will continue to do so, with the hope of heralding a new era where the clinician is better armed to combat infection and


InternalMed_Harrison

InternalMed_Harrison. prevent, and control infections has progressed rapidly in recent years and will continue to do so, with the hope of heralding a new era where the clinician is better armed to combat infection and promote human health.


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InternalMed_Harrison. Approach to the Acutely Ill Infected febrile Patient Tamar F. Barlam, Dennis L. Kasper The physician treating the acutely ill febrile patient must be able to recognize infections that require emergent attention. If such infections 147 Approach to the Acutely Ill Infected Febrile Patient are not adequately evaluated and treated at initial presentation, the opportunity to alter an adverse outcome may be lost. In this chapter, the clinical presentations of and approach to patients with relatively common infectious disease emergencies are discussed. These infectious processes and their treatments are discussed in detail in other chapters. APPROACH TO THE PATIENT:


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InternalMed_Harrison. APPROACH TO THE PATIENT: Before the history is elicited and a physical examination is performed, an immediate assessment of the patient’s general appearance can yield valuable information. The perceptive physician’s subjective sense that a patient is septic or toxic often proves accurate. Visible agitation or anxiety in a febrile patient can be a harbinger of critical illness.


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InternalMed_Harrison. HISTORY Presenting symptoms are frequently nonspecific. Detailed questions should be asked about the onset and duration of symptoms and about changes in severity or rate of progression over time. Host factors and comorbid conditions may increase the risk of infection with certain organisms or of a more fulminant course than is usually seen. Lack of splenic function, alcoholism with significant liver disease, IV drug use, HIV infection, diabetes, malignancy, organ transplantation, and chemotherapy all predispose to specific infections and frequently to increased severity. The patient should be questioned about factors that might help identify a nidus for invasive infection, such as recent upper respiratory tract infections, influenza, or varicella; prior trauma; disruption of cutaneous barriers due to lacerations, burns, surgery, body piercing, or decubiti; and the presence of foreign bodies, such as nasal packing after rhinoplasty, tampons, or prosthetic joints. Travel, contact with


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InternalMed_Harrison. due to lacerations, burns, surgery, body piercing, or decubiti; and the presence of foreign bodies, such as nasal packing after rhinoplasty, tampons, or prosthetic joints. Travel, contact with pets or other animals, or activities that might result in tick or mosquito exposure can lead to diagnoses that would not otherwise be considered. Recent dietary intake, medication use, social or occupational contact with ill individuals, vaccination history, recent sexual contacts, and menstrual history may be relevant. A review of systems should focus on any neurologic signs or sensorium alterations, rashes or skin lesions, and focal pain or tenderness and should also include a general review of respiratory, gastrointestinal, or genitourinary symptoms.


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InternalMed_Harrison. PHYSICAL EXAMINATION A complete physical examination should be performed, with special attention to several areas that are sometimes given short shrift in routine examinations. Assessment of the patient’s general appearance and vital signs, skin and soft tissue examination, and the neurologic evaluation are of particular importance. The patient may appear either anxious and agitated or lethargic and apathetic. Fever is usually present, although elderly patients and compromised hosts (e.g., patients who are uremic or cirrhotic and those who are taking glucocorticoids or nonsteroidal anti-inflammatory drugs) may be afebrile despite serious underlying infection. Measurement of blood pressure, heart rate, and respiratory rate helps determine the degree of hemodynamic and metabolic compromise. The patient’s airway must be evaluated to rule out the risk of obstruction from an invasive oropharyngeal infection.


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InternalMed_Harrison. The etiologic diagnosis may become evident in the context of a thorough skin examination (Chap. 24). Petechial rashes are typically seen with meningococcemia or Rocky Mountain spotted fever (RMSF; see Fig. 25e-16); erythroderma is associated with toxic shock syndrome (TSS) and drug fever. The soft tissue and muscle examination is critical. Areas of erythema or duskiness, edema, and tenderness may indicate underlying necrotizing fasciitis, myositis, or myonecrosis. The neurologic examination must include a careful assessment of mental status for signs of early encephalopathy. Evidence of nuchal rigidity or focal neurologic findings should be sought.


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InternalMed_Harrison. DIAGNOSTIC WORKUP After a quick clinical assessment, diagnostic material should be obtained rapidly and antibiotic and supportive treatment begun. Blood (for cultures; baseline complete blood count with differential; measurement of serum electrolytes, blood urea nitrogen, serum creatinine, and serum glucose; and liver function tests) can be obtained at the time an IV line is placed and before antibiotics are administered. The blood lactate concentration also should be measured. Three sets of blood cultures should be performed for patients with possible acute endocarditis. Asplenic patients should have a buffy coat examined for bacteria; these patients can have >106 organisms per milliliter of blood (compared with 104/mL in patients with an intact spleen). Blood smears from patients at risk for severe parasitic disease, such as malaria or babesiosis (Chap. 250e), must be examined for the diagnosis and quantitation of parasitemia. Blood smears may also be diagnostic in ehrlichiosis and


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InternalMed_Harrison. for severe parasitic disease, such as malaria or babesiosis (Chap. 250e), must be examined for the diagnosis and quantitation of parasitemia. Blood smears may also be diagnostic in ehrlichiosis and anaplasmosis.


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InternalMed_Harrison. Patients with possible meningitis should have cerebrospinal fluid (CSF) drawn before the initiation of antibiotic therapy. Focal findings, depressed mental status, or papilledema should be evaluated by brain imaging prior to lumbar puncture, which, in this setting, could initiate herniation. Antibiotics should be administered before imaging but after blood for cultures has been drawn. If CSF cultures are negative, blood cultures will provide the diagnosis in 50–70% of cases. Molecular diagnostic techniques (e.g., broad-range 16S rRNA gene polymerase chain reaction testing for bacterial meningitis pathogens) are of increasing importance in the rapid diagnosis of life-threatening infections.


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InternalMed_Harrison. Focal abscesses necessitate immediate CT or MRI as part of an evaluation for surgical intervention. Other diagnostic procedures, such as wound cultures, should not delay the initiation of treatment for more than minutes. Once emergent evaluation, diagnostic procedures, and (if appropriate) surgical consultation (see below) have been completed, other laboratory tests can be conducted. Appropriate radiography, computed axial tomography, MRI, urinalysis, erythrocyte sedimentation rate and C-reactive protein determination, and transthoracic or transesophageal echocardiography all may prove important.


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InternalMed_Harrison. In the acutely ill patient, empirical antibiotic therapy is critical and should be administered without undue delay. Increased prevalence of antibiotic resistance in community-acquired bacteria must be considered when antibiotics are selected. Table 147-1 lists first-line empirical regimens for infections considered in this chapter. In addition to the rapid initiation of antibiotic therapy, several of these infections require urgent surgical attention. Neurosurgical evaluation for subdural empyema, otolaryngologic surgery for possible mucormycosis, and cardiothoracic surgery for critically ill patients with acute endocarditis are as important as antibiotic therapy. For infections such as necrotizing fasciitis and clostridial myonecrosis, rapid surgical intervention supersedes other diagnostic or therapeutic maneuvers.


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InternalMed_Harrison. Adjunctive treatments may reduce morbidity and mortality rates and include dexamethasone for bacterial meningitis or IV immunoglobulin for TSS and necrotizing fasciitis caused by group A Streptococcus. Adjunctive therapies should usually be initiated within the first hours of treatment; however, dexamethasone for Clinical Syndrome Possible Etiologies Treatment Comments See Chap(s). Meningococcemia N. meningitidis Penicillin (4 mU q4h) or ceftriaxone Consider protein C replacement, if available, in 180 (2 g q12h) fulminant meningococcemia. Drotrecogin alfa (activated) is no longer produced. Rocky Mountain Rickettsia rickettsii Doxycycline (100 mg bid) If both meningococcemia and RMSF are being 211 spotted fever (RMSF) considered, use ceftriaxone (2 g q12h) plus doxycycline (100 mg bid). If RMSF is diagnosed, doxycycline is the proven superior agent. Purpura fulminans S. pneumoniae, H. influenzae, Ceftriaxone (2 g q12h) plus If a β-lactam-sensitive strain is identified, 171, 180,


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InternalMed_Harrison. Purpura fulminans S. pneumoniae, H. influenzae, Ceftriaxone (2 g q12h) plus If a β-lactam-sensitive strain is identified, 171, 180, N. meningitidis vancomycin (15 mg/kg q12h)b vancomycin can be discontinued. 182, 325 Erythroderma: toxic Group A Streptococcus, Vancomycin (15 mg/kg q12h)b plus If a penicillinor oxacillin-sensitive strain is 172, 173 shock syndrome Staphylococcus aureus clindamycin (600 mg q8h isolated, these agents are superior to vancomycin (penicillin, 2 mU q4h; or oxacillin, 2 g IV q4h). The site of toxigenic bacteria should be debrided; IV immunoglobulin can be used in severe cases.d Sepsis with Soft Tissue Findings Necrotizing fasciitis Group A Streptococcus, mixed Vancomycin (15 mg/kg q12h)b plus Urgent surgical evaluation is critical. Adjust 156, 172, aerobic/anaerobic flora, clindamycin (600 mg q8h) plus treatment when culture data become available. 173 CA-MRSAe gentamicin (5 mg/kg q8h)


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InternalMed_Harrison. Clostridial myonecrosis Clostridium perfringens Penicillin (2 mU q4h) plus clindamy-Urgent surgical evaluation is critical. 179 cin (600 mg q8h) Bacterial meningitis S. pneumoniae, N. meningitidis Brain abscess, Streptococcus spp., suppurative intracranial Staphylococcus spp., infections anaerobes, gram-negative Spinal epidural abscess Staphylococcus spp., gram-negative bacilli Artesunate (2.4 mg/kg IV at 0, 12, and 24 h; then once daily)f or quinine (IV loading dose of 20 mg salt/kg; then 10 mg/kg q8h) Vancomycin (15 mg/kg q12h)b plus ceftriaxone (2 g q24h) If a β-lactam-sensitive strain is identified, vancomycin can be discontinued. If the patient is >50 years old or has comorbid disease, add ampicillin (2 g q4h) for Listeria coverage. Dexamethasone (10 mg q6h × 4 days) improves outcome in adults with meningitis (especially pneumococcal) and cloudy CSF, positive CSF Gram’s stain, or a CSF leukocyte count >1000/mL.


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InternalMed_Harrison. Urgent surgical evaluation is critical. If a penicillinor oxacillin-sensitive strain is isolated, these agents are superior to vancomycin (penicillin, 4 mU q4h; or oxacillin, 2 g q4h). Do not use glucocorticoids. Use IV quinidine if IV quinine is not available. During IV quinidine treatment, blood pressure and cardiac function should be monitored continuously and blood glucose periodically. Surgical evaluation is essential. If a penicillinor oxacillin-sensitive strain is isolated, these agents are superior to vancomycin (penicillin, 4 mU q4h; or oxacillin, 2 g q4h). 246e, 248 Acute bacterial S. aureus, β-hemolytic Ceftriaxone (2 g q12h) plus Adjust treatment when culture data become 155 endocarditis streptococci, HACEK group,g vancomycin (15 mg/kg q12h)b available. Surgical evaluation is essential.


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InternalMed_Harrison. Neisseria spp., S. pneumoniae aThese empirical regimens include coverage for gram-positive pathogens that are resistant to β-lactam antibiotics. Local resistance patterns should be considered and may alter the need for empirical vancomycin. bA vancomycin loading dose of 20–25 mg/kg can be considered in critically ill patients. cβ-Lactam antibiotics may exhibit unpredictable pharmacodynamics in sepsis. Prolonged or continuous infusions can be considered. dThe optimal dose of IV immunoglobulin has not been determined, but the median dose in observational studies is 2 g/kg (total dose administered for 1–5 days). eCommunity-acquired methicillin-resistant S. aureus. fIn the United States, artesunate must be obtained through the Centers for Disease Control and Prevention. For patients diagnosed with severe malaria, full doses of parenteral antimalarial treatment should be started with whichever recommended antimalarial agent is first available. gHaemophilus species, Aggregatibacter species,


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InternalMed_Harrison. with severe malaria, full doses of parenteral antimalarial treatment should be started with whichever recommended antimalarial agent is first available. gHaemophilus species, Aggregatibacter species, Cardiobacterium hominis, Eikenella corrodens, and Kingella kingae.


InternalMed_Harrison

InternalMed_Harrison. bacterial meningitis must be given before or at the time of the first dose of antibiotic. Glucocorticoids can also be harmful, sometimes resulting in worse outcomes—e.g., when given in the setting of cerebral malaria or viral hepatitis. The infections considered below according to common clinical presentation can have rapidly catastrophic outcomes, and their immediate recognition and treatment can be life-saving. Recommended empirical therapeutic regimens are presented in Table 147-1. Patients initially have a brief prodrome of nonspecific symptoms and signs that progresses quickly to hemodynamic instability with hypo-tension, tachycardia, tachypnea, respiratory distress, and altered mental status. Disseminated intravascular coagulation (DIC) with clinical evidence of a hemorrhagic diathesis is a poor prognostic sign.


InternalMed_Harrison

InternalMed_Harrison. Septic Shock (See also Chap. 325) Patients with bacteremia leading to septic shock may have a primary site of infection (e.g., pneumonia, pyelonephritis, or cholangitis) that is not evident initially. Elderly patients with comorbid conditions, hosts compromised by malignancy and neutropenia, and patients who have recently undergone a surgical procedure or hospitalization are at increased risk for an adverse outcome. Gram-negative bacteremia with organisms such as Pseudomonas aeruginosa or Escherichia coli and gram-positive infection with organisms such as Staphylococcus aureus (including methicillin-resistant S. aureus [MRSA]) or group A streptococci can present as intractable hypotension and multiorgan failure. Treatment can usually be initiated empirically on the basis of the presentation, host factors (Chap. 325), and local patterns of bacterial resistance. Outcomes are worse when antimicrobial treatment is delayed or when the responsible pathogen ultimately proves not to be


InternalMed_Harrison

InternalMed_Harrison. host factors (Chap. 325), and local patterns of bacterial resistance. Outcomes are worse when antimicrobial treatment is delayed or when the responsible pathogen ultimately proves not to be susceptible to the initial regimen. Broad-spectrum antimicrobial agents are therefore recommended and should be instituted rapidly, preferably within the first hour after presentation. Risk factors for fungal infection should be assessed, as the incidence of fungal septic shock is increasing. Biomarkers such as C-reactive protein and procalcitonin have not proved reliable diagnostically but, when measured over time, can facilitate appropriate de-escalation of therapy. Glucocorticoids should be considered only for patients with severe sepsis who do not respond to fluid resuscitation and vasopressor therapy.


InternalMed_Harrison

InternalMed_Harrison. Overwhelming Infection in Asplenic Patients (See also Chap. 325) Patients without splenic function are at risk for overwhelming bacterial sepsis. Asplenic adult patients succumb to sepsis at 58 times the rate of the general population. Most infections are thought to occur within the first 2 years after splenectomy, with a mortality rate of ~50%, but the increased risk persists throughout life. In asplenia, encapsulated bacteria cause the majority of infections. Adults, who are more likely to have antibody to these organisms, are at lower risk than children. Streptococcus pneumoniae is the most common isolate, causing 50–70% of cases, but the risk of infection with Haemophilus influenzae or Neisseria meningitidis is also high. Severe clinical manifestations of infections due to E. coli, S. aureus, group B streptococci, P. aeruginosa, Bordetella holmesii, and Capnocytophaga, Babesia, and Plasmodium species have been described.


InternalMed_Harrison

InternalMed_Harrison. Babesiosis (See also Chap. 249) A history of recent travel to endemic areas raises the possibility of infection with Babesia. Between 1 and 4 weeks after a tick bite, the patient experiences chills, fatigue, anorexia, myalgia, arthralgia, shortness of breath, nausea, and headache; ecchymosis and/or petechiae are occasionally seen. The tick that most commonly transmits Babesia, Ixodes scapularis, also transmits Borrelia burgdorferi (the agent of Lyme disease) and Anaplasma; co-infection can occur, resulting in more severe disease. Infection with the European species Babesia divergens is more frequently fulminant than that due to the U.S. species Babesia microti. B. divergens causes a febrile syndrome with hemolysis, jaundice, hemoglobinemia, and renal failure and is associated with a mortality rate of >40%. Severe babesiosis is especially common in asplenic hosts but does occur in hosts with 781 normal splenic function, particularly those >60 years of age and those with underlying


InternalMed_Harrison

InternalMed_Harrison. rate of >40%. Severe babesiosis is especially common in asplenic hosts but does occur in hosts with 781 normal splenic function, particularly those >60 years of age and those with underlying immunosuppressive conditions such as HIV infection or malignancy. Complications include renal failure, acute respiratory failure, and DIC.


InternalMed_Harrison

InternalMed_Harrison. Other Sepsis Syndromes Tularemia (Chap. 195) is seen throughout the United States but occurs primarily in Arkansas, Missouri, South Dakota, and Oklahoma. This disease is associated with wild rabbit, tick, and tabanid fly contact. It can be transmitted by arthropod bite, handling of infected animal carcasses, consumption of contaminated food and water, or inhalation. The typhoidal form can be associated with gram-negative septic shock and a mortality rate of >30%, especially in patients with underlying comorbid or immunosuppressive conditions. Plague occurs infrequently in the United States (Chap. 196), primarily after contact with ground squirrels, prairie dogs, or chipmunks, but is endemic in other parts of the world, with >90% of all cases occurring in Africa. The septic form is particularly rare and is associated with shock, multiorgan failure, and a 30% mortality rate. These infections should be considered in the appropriate epidemiologic setting. The Centers for Disease Control


InternalMed_Harrison

InternalMed_Harrison. rare and is associated with shock, multiorgan failure, and a 30% mortality rate. These infections should be considered in the appropriate epidemiologic setting. The Centers for Disease Control and Prevention lists Francisella tularensis and Yersinia pestis (the agents of tularemia and plague, respectively) along with Bacillus anthracis (the agent of anthrax) as important organisms that might be used for bioterrorism (Chap. 261e).


InternalMed_Harrison

InternalMed_Harrison. (See also Chap. 24) Maculopapular rashes may reflect early meningococcal or rickettsial disease but are usually associated with nonemergent infections. Exanthems are usually viral. Primary HIV infection commonly presents with a rash that is typically maculopapular and involves the upper part of the body but can spread to the palms and soles. The patient is usually febrile and can have lymphadenopathy, severe headache, dysphagia, diarrhea, myalgias, and arthralgias. Recognition of this syndrome provides an opportunity to prevent transmission and to institute treatment and monitoring early on. Petechial rashes caused by viruses are seldom associated with hypotension or a toxic appearance, although there can be exceptions (e.g., severe measles or arboviral infection). Petechial rashes limited to the distribution of the superior vena cava are rarely associated with severe disease. In other settings, petechial rashes require more urgent attention.


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InternalMed_Harrison. Meningococcemia (See also Chap. 180) Almost three-quarters of patients with N. meningitidis bacteremia have a rash.


InternalMed_Harrison

InternalMed_Harrison. Meningococcemia most often affects young children (i.e., those 6 months to 5 years old). In sub-Saharan Africa, the high prevalence of serogroup A meningococcal disease has been a threat to public health for more than a century. Thousands of deaths occur annually in this area, which is known as the “meningitis belt,” and large epidemic waves occur approximately every 8–12 years. Serogroups W135 and X are also important emerging pathogens in Africa. In the United States, sporadic cases and outbreaks occur in day-care centers, schools (grade school through college, particularly among college freshmen living in residential halls), and army barracks. Household contacts of index cases are at 400–800 times greater risk of disease than the general population. Patients may exhibit fever, headache, nausea, vomiting, myalgias, changes in mental status, and meningismus. However, the rapidly progressive form of disease is not usually associated with meningitis. The rash is initially pink,


InternalMed_Harrison

InternalMed_Harrison. nausea, vomiting, myalgias, changes in mental status, and meningismus. However, the rapidly progressive form of disease is not usually associated with meningitis. The rash is initially pink, blanching, and maculopapular, appearing on the trunk and extremities, but then becomes hemorrhagic, forming petechiae. Petechiae are first seen at the ankles, wrists, axillae, mucosal surfaces, and palpebral and bulbar conjunctiva, with subsequent spread on the lower extremities and to the trunk. A cluster of petechiae may be seen at pressure points—e.g., where a blood pressure cuff has been inflated. In rapidly progressive meningococcemia (10–20% of cases), the petechial rash quickly becomes purpuric (see Fig. 70-5), and patients develop DIC, multiorgan failure, and shock; 50–60% of these patients die, and survivors often require extensive debridement or amputation of gangrenous extremities.


InternalMed_Harrison

InternalMed_Harrison. Approach to the Acutely Ill Infected Febrile Patient 782 Hypotension with petechiae for <12 h is associated with significant mortality. Cyanosis, coma, oliguria, metabolic acidosis, and elevated partial thromboplastin time also are associated with a fatal outcome. Correction of protein C deficiency may improve outcome. Antibiotics given in the office by the primary care provider before hospital evaluation and admission may improve prognosis; this observation suggests that early initiation of treatment may be life-saving. Meningococcal conjugate vaccines are protective against serogroups A, C, Y and W135 and are recommended for children 11–18 years of age and for other high-risk patients.


InternalMed_Harrison

InternalMed_Harrison. Rocky Mountain Spotted Fever (See also Chap. 211) RMSF is a tick-borne disease caused by Rickettsia rickettsii that occurs throughout North and South America. Up to 40% of patients do not report a history of a tick bite, but a history of travel or outdoor activity (e.g., camping in tick-infested areas) can often be ascertained. For the first 3 days, headache, fever, malaise, myalgias, nausea, vomiting, and anorexia are documented. By day 3, half of patients have skin findings. Blanching macules develop initially on the wrists and ankles and then spread over the legs and trunk. The lesions become hemorrhagic and are frequently petechial. The rash spreads to palms and soles later in the course. The centripetal spread is a classic feature of RMSF but occurs in a minority of patients. Moreover, 10–15% of patients with RMSF never develop a rash. The patient can be hypotensive and develop noncardiogenic pulmonary edema, confusion, lethargy, and encephalitis progressing to coma. The CSF


InternalMed_Harrison

InternalMed_Harrison. 10–15% of patients with RMSF never develop a rash. The patient can be hypotensive and develop noncardiogenic pulmonary edema, confusion, lethargy, and encephalitis progressing to coma. The CSF contains 10–100 cells/μL, usually with a predominance of mononuclear cells. The CSF glucose level is often normal; the protein concentration may be slightly elevated. Renal and hepatic injury as well as bleeding secondary to vascular damage are noted. For untreated infections, mortality rates are 20–30%. Delayed recognition and treatment are associated with a greater risk of death; Native Americans, children 5–9 years of age, adults >70 years old, and persons with underlying immunosuppression also are at increased risk of death.


InternalMed_Harrison

InternalMed_Harrison. Other rickettsial diseases cause significant morbidity and mortality worldwide. Mediterranean spotted fever caused by Rickettsia conorii is found in Africa, southwestern and south-central Asia, and southern Europe. Patients have fever, flu-like symptoms, and an inoculation eschar at the site of the tick bite. A maculopapular rash develops within 1–7 days, involving the palms and soles but sparing the face. Elderly patients or those with diabetes, alcoholism, uremia, or congestive heart failure are at risk for severe disease characterized by neurologic involvement, respiratory distress, and gangrene of the digits. Mortality rates associated with this severe form of disease approach 50%. Epidemic typhus, caused by Rickettsia prowazekii, is transmitted in louse-infested environments and emerges in conditions of extreme poverty, war, and natural disaster. Patients experience a sudden onset of high fevers, severe headache, cough, myalgias, and abdominal pain. A maculopapular rash develops


InternalMed_Harrison

InternalMed_Harrison. in conditions of extreme poverty, war, and natural disaster. Patients experience a sudden onset of high fevers, severe headache, cough, myalgias, and abdominal pain. A maculopapular rash develops (primarily on the trunk) in more than half of patients and can progress to petechiae and purpura. Serious signs include delirium, coma, seizures, noncardiogenic pulmonary edema, skin necrosis, and peripheral gangrene. Mortality rates approached 60% in the preantibiotic era and continue to exceed 10–15% in contemporary outbreaks. Scrub typhus, caused by Orientia tsutsugamushi (a separate genus in the family Rickettsiaceae), is transmitted by larval mites or chiggers and is one of the most common infections in southeastern Asia and the western Pacific. The organism is found in areas of heavy scrub vegetation (e.g., along riverbanks). Patients may have an inoculation eschar and may develop a maculopapular rash. Severe cases progress to pneumonia, meningoencephalitis, DIC, and renal failure.


InternalMed_Harrison

InternalMed_Harrison. vegetation (e.g., along riverbanks). Patients may have an inoculation eschar and may develop a maculopapular rash. Severe cases progress to pneumonia, meningoencephalitis, DIC, and renal failure. Mortality rates range from 1% to 35%.