Jason B. Harris

Characterization of dystrophin in muscle-biopsy specimens from patients with Duchenne's or Becker's muscular dystrophy.

A deficiency of the protein dystrophin has recently been shown to be the probable cause of Duchennes muscular dystrophy. We sought to determine the relation between the clinical phenotype and the status of dystrophin in muscle-biopsy specimens from 103 patients with various neuromuscular disorders. We found very low levels (less than 3 percent of normal levels) or no dystrophin in the severe Duchenne phenotype (35 of 38 patients), low concentrations of dystrophin in the intermediate (outlier) phenotype (4 of 7), and dystrophin of abnormal molecular weight in the mild Becker phenotype (12 of 18). Normal levels of dystrophin of normal molecular weight were found in nearly all the patients (38 of 40) with 20 other neuromuscular disorders we studied. These data show the clinical consequences of both quantitative alterations (in Duchennes and intermediate dystrophy) in a single protein. The biochemical assay for dystrophin should prove helpful in delineating myopathies that overlap clinically with Duchennes and Beckers dystrophies, and it shows promise as an accurate diagnostic tool.

The Origin of the Haitian Cholera Outbreak Strain

BACKGROUND Although cholera has been present in Latin America since 1991, it had not been epidemic in Haiti for at least 100 years. Recently, however, there has been a severe outbreak of cholera in Haiti. METHODS We used third-generation single-molecule real-time DNA sequencing to determine the genome sequences of 2 clinical Vibrio cholerae isolates from the current outbreak in Haiti, 1 strain that caused cholera in Latin America in 1991, and 2 strains isolated in South Asia in 2002 and 2008. Using primary sequence data, we compared the genomes of these 5 strains and a set of previously obtained partial genomic sequences of 23 diverse strains of V. cholerae to assess the likely origin of the cholera outbreak in Haiti. RESULTS Both single-nucleotide variations and the presence and structure of hypervariable chromosomal elements indicate that there is a close relationship between the Haitian isolates and variant V. cholerae El Tor O1 strains isolated in Bangladesh in 2002 and 2008. In contrast, analysis of genomic variation of the Haitian isolates reveals a more distant relationship with circulating South American isolates. CONCLUSIONS The Haitian epidemic is probably the result of the introduction, through human activity, of a V. cholerae strain from a distant geographic source. (Funded by the National Institute of Allergy and Infectious Diseases and the Howard Hughes Medical Institute.).

Cholera transmission: the host, pathogen and bacteriophage dynamic

Zimbabwe offers the most recent example of the tragedy that befalls a country and its people when cholera strikes. The 2008–2009 outbreak rapidly spread across every province and brought rates of mortality similar to those witnessed as a consequence of cholera infections a hundred years ago. In this Review we highlight the advances that will help to unravel how interactions between the host, the bacterial pathogen and the lytic bacteriophage might propel and quench cholera outbreaks in endemic settings and in emergent epidemic regions such as Zimbabwe.

Effect of Eliminating Seropositive Canines on the Transmission of Visceral Leishmaniasis in Brazil

In Brazil, where Leishmania chagasi causes endemic American visceral leishmaniasis (AVL), the spread and maintenance of human disease are attributed to canine reservoirs. However, despite measures directed toward the elimination of infected canines, the incidence of human disease continues to increase. To evaluate the role of infected canines in the acquisition of AVL by humans, we undertook a controlled intervention study in three similar, but isolated, valleys of Pancas, Espírito Santo, Brazil. In the two experimental (intervention) valleys, infected dogs were eliminated whereas in the control valley, seropositive canines remained untouched. During the 12-month study period, human seropositivity rates, as measured by dot enzyme-linked immunosorbent assay, increased from 15% to 54% in the intervention valleys and from 14% to 54% in the control valley. The elimination of infected canines in the intervention valleys did not result in a statistically significant difference between the incidences of human serological conversion in the intervention and control valleys at either 6 (20% and 22%, respectively; P = .5961) or 12 months (26% and 27%, respectively; P = .9442). The role of humans as a significant reservoir for AVL is proposed as an explanation for the study results.

Susceptibility to Vibrio cholerae Infection in a Cohort of Household Contacts of Patients with Cholera in Bangladesh

Background Despite recent progress in understanding the molecular basis of Vibrio cholerae pathogenesis, there is relatively little knowledge of the factors that determine the variability in human susceptibility to V. cholerae infection. Methods and Findings We performed an observational study of a cohort of household contacts of cholera patients in Bangladesh, and compared the baseline characteristics of household members who went on to develop culture-positive V. cholerae infection with individuals who did not develop infection. Although the vibriocidal antibody is the only previously described immunologic marker associated with protection from V. cholerae infection, we found that levels of serum IgA specific to three V. cholerae antigens—the B subunit of cholera toxin, lipopolysaccharide, and TcpA, the major component of the toxin–co-regulated pilus—also predicted protection in household contacts of patients infected with V. cholerae O1, the current predominant cause of cholera. Circulating IgA antibodies to TcpA were also associated with protection from V. cholerae O139 infection. In contrast, there was no association between serum IgG antibodies specific to these three antigens and protection from infection with either serogroup. We also found evidence that host genetic characteristics and serum retinol levels modify susceptibility to V. cholerae infection. Conclusions Our observation that levels of serum IgA (but not serum IgG) directed at certain V. cholerae antigens are associated with protection from infection underscores the need to better understand anti–V. cholerae immunity at the mucosal surface. Furthermore, our data suggest that susceptibility to V. cholerae infection is determined by a combination of immunologic, nutritional, and genetic characteristics; additional factors that influence susceptibility to cholera remain unidentified.

Blood Group, Immunity, and Risk of Infection with Vibrio cholerae in an Area of Endemicity

ABSTRACT Individuals with blood group O are more susceptible than other individuals to severe cholera, although the mechanism underlying this association is unknown. To assess the respective roles of both intrinsic host factors and adaptive immune responses that might influence susceptibility to infection with Vibrio cholerae, we prospectively followed a cohort of household contacts of patients with cholera in Bangladesh. In this study, we made the novel observation that persons with blood group O were less likely than those with other blood groups to become infected with V. cholerae O1 (odds ratio [OR], 0.67; 95% confidence interval [CI], 0.53 to 0.85; P = 0.008). Consistent with prior studies, however, household contacts with blood group O were more likely to develop severe illness if infected with V. cholerae O1 (OR, 2.3; 95% CI, 0.98 to 5.59; P = 0.05). While blood group O protected significantly against infection with V. cholerae O1, there was no evidence of protection against V. cholerae O139. A multivariate analysis demonstrated that the association between blood group O and protection from infection with V. cholerae O1 was independent of age, gender, and baseline anti-cholera toxin and vibriocidal antibody titers. Based on this epidemiologic evidence, we propose a hypothesis for understanding the association between blood group O and the risk of infection with V. cholerae O1 and O139 as well as the risk of developing severe symptoms once infected.

Meeting cholera's challenge to Haiti and the world: a joint statement on cholera prevention and care.

Cholera in Haiti: Acute-on-Chronic Long before the devastating earthquake on January 12, 2010, Haiti struggled beneath the burdens of intractable poverty and ill health. The poorest country in the Western Hemisphere, Haiti also faces some of the highest rates of maternal and infant mortality—widely used indicators of the robustness of a health system—in the world ([S1] in Text S1; [2], [3]). The October 2010 cholera outbreak is the most recent of a long series of affronts to the health of Haitis population; it is yet another acute symptom of the chronic weakness of Haitis health, water, and sanitation systems. Water and sanitation conditions highlight these systemic weaknesses. In 2002, Haiti ranked last out of 147 countries for water security [4], [5]. Before the earthquake struck, only half of the population in the capital, Port-au-Prince, had access to latrines or other forms of modern sanitation, and roughly one-third had no access to tap water [6]. Across the country, access to sanitation and clean water is even more limited: only 17% of Haitians had access to adequate sanitation in 2008, and 12% received treated water [7]. Not surprisingly, diarrheal diseases have long been a significant cause of death and disability, especially among children under 5 years of age [6]. The cholera outbreak began less than a year after a 7.0-magnitude earthquake took the lives of more than 300,000 people and left nearly 1.5 million homeless [6]. Almost 1 million Haitians still live in spontaneous settlements known as internally displaced persons (IDP) camps [8]. While post-earthquake conditions in Haiti were ripe for outbreaks of acute diarrheal illness, cholera was deemed “very unlikely to occur” by the United States Centers for Disease Control and Prevention (CDC) and other public health authorities [9]. Cholera had never before been reported in Haiti [S2] [10], [11]; health providers were unprepared for an influx of patients presenting with acute watery diarrhea. The cholera epidemic has been most severe in rural areas and large urban slums. Rural communities were charged with hosting hundreds of thousands of displaced people after the earthquake, placing greater demands on their already-scarce resources, including water. Surface water drawn directly from the source or piped from rivers and streams constitutes the principal supply of drinking water in rural Haiti. The lack of adequate piping, filtration, and water treatment systems (including chlorination) made these rural regions vulnerable to the rapid spread of waterborne disease. While most IDP camps have been supplied with potable water, large urban slums have had to rely on existing water sources—some of them containing Vibrio cholerae—and have therefore been vulnerable to rapid disease spread. Most slums also have poor sanitation infrastructure. Since the first cases were reported in Saint-Marc and Mirebalais, cholera has spread to every department in Haiti, and to other countries, too [S3] [12]–[14]. Public suspicion (ultimately validated by genomic sequence analyses [15]) of the strains link to South Asia, home to a group of United Nations peacekeepers stationed in central Haiti, triggered blame and violence that interfered with response efforts. As we have learned from the global AIDS pandemic and other infectious disease epidemics, cycles of accusation can continue for years, diverting attention and resources from the delivery of care and prevention services [16]. Systemic problems that brought cholera to epidemic levels in Haiti will (unless addressed) continue to facilitate its spread. As a disease of poverty, cholera preys upon the bottom of the social gradient; international trade, migration, and travel—from South Asia or elsewhere—open direct channels for pathogens that follow social fault lines.

Transcriptional Profiling of Vibrio cholerae Recovered Directly from Patient Specimens during Early and Late Stages of Human Infection

ABSTRACT Understanding gene expression by bacteria during the actual course of human infection may provide important insights into microbial pathogenesis. In this study, we evaluated the transcriptional profile of Vibrio cholerae, the causative agent of cholera, in clinical specimens from cholera patients. We collected samples of human stool and vomitus that were positive by dark-field microscopy for abundant vibrios and used a microarray to compare gene expression in organisms recovered directly from specimens collected during the early and late stages of human infection. Our results reveal that V. cholerae gene expression within the human host environment differs from patterns defined in in vitro models of pathogenesis. tcpA, the major subunit of the essential V. cholerae colonization factor, was significantly more highly expressed in early than in late stages of infection; however, the genes encoding cholera toxin were not highly expressed in either phase of human infection. Furthermore, expression of the virulence regulators toxRS and tcpPH was uncoupled. Interestingly, the pattern of gene expression indicates that the human upper intestine may be a uniquely suitable environment for the transfer of genetic elements that are important in the evolution of pathogenic strains of V. cholerae. These findings provide a more detailed assessment of the transcriptome of V. cholerae in the human host than previous studies of organisms in stool alone and have implications for cholera control and the design of improved vaccines.

Antigen-Specific Memory B-Cell Responses to Vibrio cholerae O1 Infection in Bangladesh

ABSTRACT Cholera, caused by Vibrio cholerae, is a noninvasive dehydrating enteric disease with a high mortality rate if untreated. Infection with V. cholerae elicits long-term protection against subsequent disease in countries where the disease is endemic. Although the mechanism of this protective immunity is unknown, it has been hypothesized that a protective mucosal response to V. cholerae infection may be mediated by anamnestic responses of memory B cells in the gut-associated lymphoid tissue. To characterize memory B-cell responses to cholera, we enrolled a cohort of 39 hospitalized patients with culture-confirmed cholera and evaluated their immunologic responses at frequent intervals over the subsequent 1 year. Memory B cells to cholera antigens, including lipopolysaccharide (LPS), and the protein antigens cholera toxin B subunit (CTB) and toxin-coregulated pilus major subunit A (TcpA) were enumerated using a method of polyclonal stimulation of peripheral blood mononuclear cells followed by a standard enzyme-linked immunospot procedure. All patients demonstrated CTB, TcpA, and LPS-specific immunoglobulin G (IgG)and IgA memory responses by day 90. In addition, these memory B-cell responses persisted up to 1 year, substantially longer than other traditional immunologic markers of infection with V. cholerae. While the magnitude of the LPS-specific IgG memory B-cell response waned at 1 year, CTB- and TcpA-specific IgG memory B cells remained significantly elevated at 1 year after infection, suggesting that T-cell help may result in a more durable memory B-cell response to V. cholerae protein antigens. Such memory B cells could mediate anamnestic responses on reexposure to V. cholerae.

Clinical Outcomes in Household Contacts of Patients with Cholera in Bangladesh

BACKGROUND Multiple Vibrio cholerae infections in the same household are common. The objective of this study was to examine the incidence of V. cholerae infection and associated clinical symptoms in household contacts of patients with cholera and to identify risk factors for development of severe dehydration in this cohort. METHODS Household contacts of hospitalized patients with cholera were observed with frequent clinical assessments and collection of serum and rectal swab samples for culture for a period of 21 days after presentation of the index case. RESULTS One-half (460 of 944) of all contacts reported diarrhea during the study period, and symptoms most frequently began 2 days after presentation of the index case. Antibiotics were used by 199 (43%) of 460 contacts with diarrhea. Results of rectal swab cultures for V. cholerae were positive for 202 (21%) of 944 contacts, and 148 (73%) infected contacts experienced diarrhea. Significant dehydration developed in 26 contacts; predictors of dehydration included vomiting, each additional day of diarrhea, and blood group O status. CONCLUSIONS In urban Bangladesh, the burden of diarrheal illness among household contacts of patients with cholera is higher than was previously estimated, and prophylactic intervention is feasible, because the majority of symptomatic cases of V. cholerae infection in contacts begin soon after presentation of the index case. Re-evaluation of targeted chemoprophylaxis for household contacts of patients with cholera may be warranted.