[Show abstract][Hide abstract] ABSTRACT: During outbreaks of infectious diseases or in cases of severely ill patients, it is imperative to identify the causative agent. This report describes several events in which virus isolation and identification by electron microscopy were critical to initial recognition of the etiologic agent, which was further analyzed by additional laboratory diagnostic assays. Examples include severe acute respiratory syndrome coronavirus, and Nipah, lymphocytic choriomeningitis, West Nile, Cache Valley, and Heartland viruses. These cases illustrate the importance of the techniques of cell culture and electron microscopy in pathogen identification and recognition of emerging diseases.
[Show abstract][Hide abstract] ABSTRACT: Marburg virus (family Filoviridae) causes sporadic outbreaks of severe hemorrhagic disease in sub-Saharan Africa. Bats have been implicated as likely natural reservoir hosts based most recently on an investigation of cases among miners infected in 2007 at the Kitaka mine, Uganda, which contained a large population of Marburg virus-infected Rousettus aegyptiacus fruit bats. Described here is an ecologic investigation of Python Cave, Uganda, where an American and a Dutch tourist acquired Marburg virus infection in December 2007 and July 2008. More than 40,000 R. aegyptiacus were found in the cave and were the sole bat species present. Between August 2008 and November 2009, 1,622 bats were captured and tested for Marburg virus. Q-RT-PCR analysis of bat liver/spleen tissues indicated ∼2.5% of the bats were actively infected, seven of which yielded Marburg virus isolates. Moreover, Q-RT-PCR-positive lung, kidney, colon and reproductive tissues were found, consistent with potential for oral, urine, fecal or sexual transmission. The combined data for R. aegyptiacus tested from Python Cave and Kitaka mine indicate low level horizontal transmission throughout the year. However, Q-RT-PCR data show distinct pulses of virus infection in older juvenile bats (∼six months of age) that temporarily coincide with the peak twice-yearly birthing seasons. Retrospective analysis of historical human infections suspected to have been the result of discrete spillover events directly from nature found 83% (54/65) events occurred during these seasonal pulses in virus circulation, perhaps demonstrating periods of increased risk of human infection. The discovery of two tags at Python Cave from bats marked at Kitaka mine, together with the close genetic linkages evident between viruses detected in geographically distant locations, are consistent with R. aegyptiacus bats existing as a large meta-population with associated virus circulation over broad geographic ranges. These findings provide a basis for developing Marburg hemorrhagic fever risk reduction strategies.
[Show abstract][Hide abstract] ABSTRACT: Nipah virus (NiV) is a highly pathogenic paramyxovirus that causes fatal encephalitis in humans. The initial outbreak of NiV infection occurred in Malaysia and Singapore in 1998-1999; relatively small, sporadic outbreaks among humans have occurred in Bangladesh since 2001. We characterized the complete genomic sequences of identical NiV isolates from 2 patients in 2008 and partial genomic sequences of throat swab samples from 3 patients in 2010, all from Bangladesh. All sequences from patients in Bangladesh comprised a distinct genetic group. However, the detection of 3 genetically distinct sequences from patients in the districts of Faridpur and Gopalganj indicated multiple co-circulating lineages in a localized region over a short time (January-March 2010). Sequence comparisons between the open reading frames of all available NiV genes led us to propose a standardized protocol for genotyping NiV; this protcol provides a simple and accurate way to classify current and future NiV sequences.
[Show abstract][Hide abstract] ABSTRACT: Alkhurma hemorrhagic fever virus (AHFV) and Kyasanur forest disease virus (KFDV) cause significant human disease and mortality in Saudi Arabia and India, respectively. Despite their distinct geographic ranges, AHFV and KFDV share a remarkably high sequence identity. Given its emergence decades after KFDV, AHFV has since been considered a variant of KFDV and thought to have arisen from an introduction of KFDV to Saudi Arabia from India. To gain a better understanding of the evolutionary history of AHFV and KFDV, we analyzed the full length genomes of 16 AHFV and 3 KFDV isolates.
Viral genomes were sequenced and compared to two AHFV sequences available in GenBank. Sequence analyses revealed higher genetic diversity within AHFVs isolated from ticks than human AHFV isolates. A Bayesian coalescent phylogenetic analysis demonstrated an ancient divergence of AHFV and KFDV of approximately 700 years ago.
The high sequence diversity within tick populations and the presence of competent tick vectors in the surrounding regions, coupled with the recent identification of AHFV in Egypt, indicate possible viral range expansion or a larger geographic range than previously thought. The divergence of AHFV from KFDV nearly 700 years ago suggests other AHFV/KFDV-like viruses might exist in the regions between Saudi Arabia and India. Given the human morbidity and mortality associated with these viruses, these results emphasize the importance of more focused study of these significant public health threats.
[Show abstract][Hide abstract] ABSTRACT: We investigated a cluster of patients with encephalitis in the Manikgonj and Rajbari Districts of Bangladesh in February 2008 to determine the etiology and risk factors for disease.
We classified persons as confirmed Nipah cases by the presence of immunoglobulin M antibodies against Nipah virus (NiV), or by the presence of NiV RNA or by isolation of NiV from cerebrospinal fluid or throat swabs who had onset of symptoms between February 6 and March 10, 2008. We classified persons as probable cases if they reported fever with convulsions or altered mental status, who resided in the outbreak areas during that period, and who died before serum samples were collected. For the case-control study, we compared both confirmed and probable Nipah case-patients to controls, who were free from illness during the reference period. We used motion-sensor-infrared cameras to observe bat's contact of date palm sap.
We identified four confirmed and six probable case-patients, nine (90%) of whom died. The median age of the cases was 10 years; eight were males. The outbreak occurred simultaneously in two communities that were 44 km apart and separated by a river. Drinking raw date palm sap 2-12 days before illness onset was the only risk factor most strongly associated with the illness (adjusted odds ratio 25, 95% confidence intervals 3.3-∞, p<0.001). Case-patients reported no history of physical contact with bats, though community members often reported seeing bats. Infrared camera photographs showed that Pteropus bats frequently visited date palm trees in those communities where sap was collected for human consumption.
This is the second Nipah outbreak in Bangladesh where date palm sap has been implicated as the vehicle of transmission. Fresh date palm sap should not be drunk, unless effective steps have been taken to prevent bat access to the sap during collection.
[Show abstract][Hide abstract] ABSTRACT: Hantaviruses are distributed throughout the United States and are the etiologic agents for hantavirus pulmonary syndrome and hemorrhagic fever with renal syndrome. Hantavirus genotypes and epidemiologic patterns vary spatially across the United States. While several longitudinal studies have been performed in the western United States, little is known about the virus in the eastern United States. We undertook a longitudinal study of hantaviruses in the primary rodent reservoir host in central Pennsylvania, Peromyscus leucopus. Prevalence of hantavirus antibodies varied both by year and site, but was not correlated with host abundance. Males were significantly more likely to have antibodies to a hantavirus than females, and both antibody sero-conversion and antibody prevalence increased with mass class (indicator for age). Our findings suggest that one or more hantaviruses are present and circulating among P. leucopus of central Pennsylvania, and understanding the dynamics in this region could help prevent zoonotic transmission to humans. Our aim was to describe the differences in epizootiology of hantavirus infection in rodents from various geographical locations to enable improved analysis of the risk of rodent-to-human transmission and obtain insights that may indicate improved means of disease intervention.
[Show abstract][Hide abstract] ABSTRACT: In March 2007, we investigated a cluster of Nipah encephalitis to identify risk factors for Nipah infection in Bangladesh.
We defined confirmed Nipah cases by the presence of IgM and IgG antibodies against Nipah virus in serum. Case-patients, who resided in the same village during the outbreak period but died before serum could be collected, were classified as probable cases.
We identified three confirmed and five probable Nipah cases. There was a single index case. Five of the secondary cases came in close physical contact to the index case when she was ill. Case-patients were more likely to have physical contact with the index case (71% cases versus 0% controls, p = <0.001). The index case, on her third day of illness, and all the subsequent cases attended the same religious gathering. For three probable cases including the index case, we could not identify any known risk factors for Nipah infection such as physical contact with Nipah case-patients, consumption of raw date palm juice, or contact with sick animals or fruit bats.
Though person-to-person transmission remains an important mode of transmission for Nipah infection, we could not confirm the source of infection for three of the probable Nipah case-patients. Continued surveillance and outbreak investigations will help better understand the transmission of Nipah virus and develop preventive strategies.
PLoS ONE 10/2010; 5(10):e13570. DOI:10.1371/journal.pone.0013570 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Sin Nombre virus (SNV) is the primary cause of hantavirus pulmonary syndrome (HPS) in the United States. Although other studies have demonstrated a possible association between neutralizing antibody titers and the severity of HPS, the exact nature of serologic responses and their association with outcomes have not been fully characterized.
We examined immunoglobulin M (IgM) and immunoglobulin G (IgG) serologic responses in 94 clinical samples from 81 patients with confirmed HPS. We further compared a subset of 31 patients with fatal HPS and 20 surviving patients for whom samples were available within a week after the onset of HPS.
SNV-specific IgM antibodies displayed a trend suggesting an early peak, whereas IgG antibody values peaked later. Among individuals with samples from the first week after the onset of HPS, all surviving patients had SNV-specific IgG responses, compared with <50% of patients with fatal HPS, and the distribution of IgG responses was significantly higher in surviving patients.
Production of SNV-specific IgM antibodies occurs early during the clinical course of HPS, whereas production of IgG antibodies may be more protracted. The presence and overall distribution of higher IgG antibody titers in surviving patients with HPS suggests that production of SNV-specific IgG may be a strong predictor of favorable outcomes.
The Journal of Infectious Diseases 07/2010; 202(2):242-6. DOI:10.1086/653482 · 6.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Lymphocytic choriomeningitis virus (LCMV) is the prototype of the family Arenaviridae. LCMV can be associated with severe disease in humans, and its global distribution reflects the broad dispersion of the primary rodent reservoir, the house mouse (Mus musculus). Recent interest in the natural history of the virus has been stimulated by increasing recognition of LCMV infections during pregnancy, and in clusters of LCMV-associated fatal illness among tissue transplant recipients. Despite its public health importance, little is known regarding the genetic diversity or distribution of virus variants. Genomic analysis of 29 LCMV strains collected from a variety of geographic and temporal sources showed these viruses to be highly diverse. Several distinct lineages exist, but there is little correlation with time or place of isolation. Bayesian analysis estimates the most recent common ancestor to be 1,000-5,000 years old, and this long history is consistent with complex phylogeographic relationships of the extant virus isolates.
[Show abstract][Hide abstract] ABSTRACT: Human Nipah outbreaks recur in a specific region and time of year in Bangladesh. Fruit bats are the reservoir host for Nipah virus. We identified 23 introductions of Nipah virus into human populations in central and northwestern Bangladesh from 2001 through 2007. Ten introductions affected multiple persons (median 10). Illness onset occurred from December through May but not every year. We identified 122 cases of human Nipah infection. The mean age of case-patients was 27 years; 87 (71%) died. In 62 (51%) Nipah virus-infected patients, illness developed 5-15 days after close contact with another Nipah case-patient. Nine (7%) Nipah case-patients transmitted virus to others. Nipah case-patients who had difficulty breathing were more likely than those without respiratory difficulty to transmit Nipah (12% vs. 0%, p = 0.03). Although a small minority of infected patients transmit Nipah virus, more than half of identified cases result from person-to-person transmission. Interventions to prevent virus transmission from bats to humans and from person to person are needed.
[Show abstract][Hide abstract] ABSTRACT: Author Summary
Marburg virus, similar to its close cousin Ebola virus, can cause large outbreaks of hemorrhagic fever (HF) in rural Africa with case fatalities approaching 90%. For decades, a long-standing enigma has been the identity of the natural reservoir of this deadly virus. In this report, we identify the cave-dwelling Egyptian fruit bat (Rousettus aegyptiacus) as a natural host of Marburg virus based on multiple lines of evidence which include, for the first time ever, the isolation of virus directly from wild-caught and apparently healthy bats. The species R. aegyptiacus is common throughout Africa with distribution into the eastern Mediterranean and Middle East. Our finding of active virus infection in approximately 5% of R. aegyptiacus bats and their population exceeding 100,000 in Kitaka cave in Uganda suggests there are likely over 5,000 Marburg virus–infected bats in this cave, which is only one of many such cave populations throughout Africa. Clearly, these bats could serve as a major source of virus with potential to initiate human epidemics, and the implications for public health are striking. Additionally, we found highly divergent (21%) genome sequences among viruses circulating in these bat populations, a level of diversity that would result from a long-term association with a suitable reservoir host of large population size.
[Show abstract][Hide abstract] ABSTRACT: Etiologic studies of acute febrile disease were conducted in sites across South America, including Cusco and Iquitos, Peru. Patients' clinical signs and symptoms were recorded, and acute- and convalescent-phase serum samples were obtained for serologic examination and virus isolation in Vero E6 and C6/36 cells. Virus isolated in Vero E6 cells was identified as encephalomyocarditis virus (EMCV) by electron microscopy and by subsequent molecular diagnostic testing of samples from 2 febrile patients with nausea, headache, and dyspnea. The virus was recovered from acute-phase serum samples from both case-patients and identified with cardiovirus-specific reverse transcription-PCR and sequencing. Serum samples from case-patient 1 showed cardiovirus antibody by immunoglobulin M ELISA (acute phase <8, convalescent phase >1,024) and by neutralization assay (acute phase <10, convalescent phase >1,280). Serum samples from case-patient 2 did not contain antibodies detectable by either assay. Detection of virus in serum strongly supports a role for EMCV in human infection and febrile illness.
[Show abstract][Hide abstract] ABSTRACT: Author Summary
In this report we describe a newly discovered ebolavirus species which caused a large hemorrhagic fever outbreak in western Uganda. The virus is genetically distinct, differing by more than 30% at the genome level from all other known ebolavirus species. The unique nature of this virus created challenges for traditional filovirus molecular based diagnostic assays and genome sequencing approaches. Instead, we quickly determined over 70% of the virus genome using a recently developed random-primed pyrosequencing approach that allowed the rapid development of a molecular detection assay that was deployed in the disease outbreak response. This draft sequence allowed easy completion of the whole genome sequence using a traditional primer walking approach and prompt confirmation that this virus represented a new ebolavirus species. Current efforts to design effective diagnostics, antivirals and vaccines will need to take into account the distinct nature of this important new member of the filovirus family.
[Show abstract][Hide abstract] ABSTRACT: Rift Valley fever (RVF) virus historically has caused widespread and extensive outbreaks of severe human and livestock disease throughout Africa, Madagascar, and the Arabian Peninsula. Following unusually heavy rainfall during the late autumn of 2006, reports of human and animal illness consistent with RVF virus infection emerged across semiarid regions of the Garissa District of northeastern Kenya and southern Somalia. Following initial RVF virus laboratory confirmation, a high-throughput RVF diagnostic facility was established at the Kenyan Central Veterinary Laboratories in Kabete, Kenya, to support the real-time identification of infected livestock and to facilitate outbreak response and control activities. A total of 3,250 specimens from a variety of animal species, including domesticated livestock (cattle, sheep, goats, and camels) and wildlife collected from a total of 55 of 71 Kenyan administrative districts, were tested by molecular and serologic assays. Evidence of RVF infection was found in 9.2% of animals tested and across 23 districts of Kenya, reflecting the large number of affected livestock and the geographic extent of the outbreak. The complete S, M, and/or L genome segment sequence was obtained from a total of 31 RVF virus specimens spanning the entire known outbreak period (December-May) and geographic areas affected by RVF virus activity. Extensive genomic analyses demonstrated the concurrent circulation of multiple virus lineages, gene segment reassortment, and the common ancestry of the 2006/2007 outbreak viruses with those from the 1997-1998 east African RVF outbreak. Evidence of recent increases in genomic diversity and effective population size 2 to 4 years prior to the 2006-2007 outbreak also was found, indicating ongoing RVF virus activity and evolution during the interepizootic/epidemic period. These findings have implications for further studies of basic RVF virus ecology and the design of future surveillance/diagnostic activities, and they highlight the critical need for safe and effective vaccines and antiviral compounds to combat this significant veterinary and public health threat.
Journal of Virology 10/2008; 82(22):11152-66. DOI:10.1128/JVI.01519-08 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Author Summary
Four rodent-borne arenaviruses are known to cause hemorrhagic fever (HF) in the New World. These include Junín, Machupo, Guanarito, and Sabiá viruses, which are found in rural areas of Argentina, Bolivia, Venezuela, and Brazil, respectively. In December 2003 and January 2004, a small number of HF cases were reported in rural Bolivia in an area outside the known Machupo HF endemic zone, and sera from one fatal case was available for laboratory testing. The man had symptoms similar to those seen with other arenaviral HF cases—acute febrile illness beginning with headache, joint and muscle pain, and vomiting—and rapidly progressed to shock, bleeding, and death at 14 days post onset of illness. Virus was isolated from two of the patient's serum samples and identified as an arenavirus by reaction of virus infected cells with arenavirus-specific antibodies and by genetic detection techniques (PCR). Subsequent complete genome analysis of the virus showed the virus to be a distinct newly discovered member of the arenavirus family, and the name Chapare virus was proposed. The virus is phylogenetically related to other arenaviruses that naturally cause hemorrhagic fever in South America, particularly Sabiá virus. Physicians should consider Chapare virus as a potential etiologic agent when encountering HF cases in the region.
[Show abstract][Hide abstract] ABSTRACT: In Bangladesh, 4 outbreaks of Nipah virus infection were identified during the period 2001-2004.
We characterized the clinical features of Nipah virus-infected individuals affected by these outbreaks. We classified patients as having confirmed cases of Nipah virus infection if they had antibodies reactive with Nipah virus antigen. Patients were considered to have probable cases of Nipah virus infection if they had symptoms consistent with Nipah virus infection during the same time and in the same community as patients with confirmed cases.
We identified 92 patients with Nipah virus infection, 67 (73%) of whom died. Although all age groups were affected, 2 outbreaks principally affected young persons (median age, 12 years); 62% of the affected persons were male. Fever, altered mental status, headache, cough, respiratory difficulty, vomiting, and convulsions were the most common signs and symptoms; clinical and radiographic features of acute respiratory distress syndrome of Nipah illness were identified during the fourth outbreak. Among those who died, death occurred a median of 6 days (range, 2-36 days) after the onset of illness. Patients who died were more likely than survivors to have a temperature >37.8 degrees C, altered mental status, difficulty breathing, and abnormal plantar reflexes. Among patients with Nipah virus infection who had well-defined exposure to another patient infected with Nipah virus, the median incubation period was 9 days (range, 6-11 days).
Nipah virus infection produced rapidly progressive severe illness affecting the central nervous and respiratory systems. Clinical characteristics of Nipah virus infection in Bangladesh, including a severe respiratory component, appear distinct from clinical characteristics reported during earlier outbreaks in other countries.
[Show abstract][Hide abstract] ABSTRACT: An encephalitis outbreak was investigated in Faridpur District, Bangladesh, in April-May 2004 to determine the cause of the outbreak and risk factors for disease. Biologic specimens were tested for Nipah virus. Surfaces were evaluated for Nipah virus contamination by using reverse transcription-PCR (RT-PCR). Thirty-six cases of Nipah virus illness were identified; 75% of case-patients died. Multiple peaks of illness occurred, and 33 case-patients had close contact with another Nipah virus patient before their illness. Results from a case-control study showed that contact with 1 patient carried the highest risk for infection (odds ratio 6.7, 95% confidence interval 2.9-16.8, p < 0.001). RT-PCR testing of environmental samples confirmed Nipah virus contamination of hospital surfaces. This investigation provides evidence for person-to-person transmission of Nipah virus. Capacity for person-to-person transmission increases the potential for wider spread of this highly lethal pathogen and highlights the need for infection control strategies for resource-poor settings.
[Show abstract][Hide abstract] ABSTRACT: We conducted a seroprevalence study and exposure survey of healthcare workers to assess the risk of nosocomial transmission of Nipah virus during an outbreak in Bangladesh in 2004. No evidence of recent Nipah virus infection was detected despite substantial exposures and minimal use of personal protective equipment.
Infection Control and Hospital Epidemiology 07/2007; 28(6):740-2. DOI:10.1086/516665 · 4.18 Impact Factor