ACKGROUND

Critically ill patients have considerable oxidative stress. Glutamine and antioxidant supplementation may offer therapeutic benefit, although current data are conflicting.

METHODS
In this blinded 2-by-2 factorial trial, we randomly assigned 1223 critically ill adults in 40 intensive care units (ICUs) in Canada, the United States, and Europe who had multiorgan failure and were receiving mechanical ventilation to receive supplements of glutamine, antioxidants, both, or placebo. Supplements were started within 24 hours after admission to the ICU and were provided both intravenously and enterally. The primary outcome was 28-day mortality. Because of the interim-analysis plan, a P value of less than 0.044 at the final analysis was considered to indicate statistical significance.

RESULTS
There was a trend toward increased mortality at 28 days among patients who received glutamine as compared with those who did not receive glutamine (32.4% vs. 27.2%; adjusted odds ratio, 1.28; 95% confidence interval [CI], 1.00 to 1.64; P=0.05). In-hospital mortality and mortality at 6 months were significantly higher among those who received glutamine than among those who did not. Glutamine had no effect on rates of organ failure or infectious complications. Antioxidants had no effect on 28-day mortality (30.8%, vs. 28.8% with no antioxidants; adjusted odds ratio, 1.09; 95% CI, 0.86 to 1.40; P=0.48) or any other secondary end point. There were no differences among the groups with respect to serious adverse events (P=0.83).

CONCLUSIONS
Early provision of glutamine or antioxidants did not improve clinical outcomes, and glutamine was associated with an increase in mortality among critically ill patients with multiorgan failure. (Funded by the Canadian Institutes of Health Research; ClinicalTrials.gov number, NCT00133978.) Sporadic human infections with avian influenza A viruses, which usually occur after recent exposure to poultry, have caused a wide spectrum of illness, ranging from conjunctivitis and upper respiratory tract disease to pneumonia and multiorgan failure. Low pathogenic avian influenza A (H7N2, H7N3, H9N2, or H10N7)1-4 virus infections have caused lower respiratory tract illness that is mild (conjunctivitis or uncomplicated influenza-like illness) to moderate in severity. Most human infections with highly pathogenic avian influenza (HPAI) A (H7) viruses have resulted in conjunctivitis (H7N3) or uncomplicated influenza illness, but one case of fatal acute respiratory distress syndrome (ARDS) was reported in a patient with H7N7 virus infection during an outbreak in the Netherlands.1,5In contrast, the cumulative case fatality rate since 2003 for reported cases of HPAI H5N1 virus infection is approximately 60%. 6-8 The transmission of H7 viruses to mammals has been reported only rarely9 in Asia. Human infections with N9 subtype viruses had not been documented anywhere in the world. In February and March 2013, three patients were hospitalized with severe lower respiratory tract disease of unknown cause. We report the identification of a novel avian-origin reassortant influenza A (H7N9) virus associated with these infections.

METHODS
Surveillance, Reporting, and Data Collection

Throat-swab specimens obtained from three adult Chinese patients (two from Shanghai City and one from Anhui Province) who were hospitalized with severe bilateral pneumonia, leukopenia, and lymphocytopenia were sent to Shanghai Public Health Clinical Center, the Shanghai Centers for Disease Control and Prevention (CDC), and the Anhui CDC, respectively. After preliminary detection of respiratory pathogens, the samples were sent to the Chinese National Influenza Center (CNIC) on March 25, 2013. A standardized surveillance reporting form was used to collect epidemiologic and clinical data, including demographic characteristics; underlying medical conditions; history of seasonal influenza vaccination; recent exposures to swine, poultry, or other animals; recent visits to a live animal market; clinical signs and symptoms; chest radiographic findings; laboratory testing results, including diagnostic testing for influenza and other respiratory viruses; antiviral treatment; clinical complications; and outcomes. A confirmed case of human infection with avian-origin influenza A (H7N9) virus was defined as evidence of pneumonia with H7N9 viral RNA or isolation of H7N9 virus from respiratory specimens at the CNIC. Isolation of the Virus Throat-swab specimens obtained from all three patients were maintained in a viral-transport medium. The specimens were propagated in the allantoic sac and amniotic cavity of 9-to-11-day-old specific pathogenfree embryonated chicken eggs for 48 to 72 hours at 35C. RNA Extraction and Real-Time RT-PCR RNA was extracted from throat-swab samples with the use of the QIAamp Viral RNA Mini Kit (Qiagen), according to the manufacturer's instructions. Specific real-time reverse-transcriptasepolymerase-chainreaction (RT-PCR) assays for seasonal influenza viruses (H1, H3, or B), H5N1, severe acute respiratory syndrome coronavirus (SARS-CoV), and novel coronavirus were used. Real-time RT-PCR assays with selfdesigned specific primer and probe sets for detecting H1 to H16 and N1 to N9 subtypes were then performed to verify the viral subtypes. Genome Sequencing and Phylogenetic Analysis A total of 198 primer sets were used to amplify the full genome for sequencing, with the use of Qiagen OneStep RT-PCR Kit. PCR products were purified from agarose gel with the use of the QIAquick Gel Extraction Kit (Qiagen). We performed the sequencing using an ABI 3730xl automatic DNA analyzer (Life Technologies) and the ABI BigDye Terminator v3.1 cycle sequencing kit (Life Technologies), according to the manufacturer's recommendations. Full genome sequences of the viruses from these patients were deposited in the Global Initiative on Sharing Avian Influenza Data (GISAID) database on March 29, 2013 (accession numbers are provided in Table S1 in theSupplementary Appendix, available with the full text of this article at NEJM.org). We performed multiple sequence alignments with the ClustalW program using MEGA software, version 5.05. Phylogenetic trees were constructed by means of the neighbor-joining method with the use of MEGA software, version 5.05, to estimate the viral gene relationship with selected influenza A virus strains obtained from GenBank.

RESULTS
Patients Patient 1 was an 87-year-old man with chronic obstructive pulmonary disease (COPD) and hypertension who reported a cough and sputum production at the onset of illness. High fever and dyspnea developed 1

week after the onset of illness. He had no known history of exposure to live birds during the 2 weeks before the onset of symptoms. Patient 2 was a 27-year-old man with a history of hepatitis B virus infection with positive hepatitis B surface antigen who presented to the hospital with high fever and cough. This patient was a butcher who worked at a market where there were transactions involving live birds. He sold pork but had not butchered bird meat before the onset of illness. Both Patient 1 and Patient 2 lived in Min-hang District, Shanghai, and were admitted to the Fifth People's Hospital. Patient 3 was a 35-year-old woman who lived in Anhui Province. She had a history of depression, hepatitis B virus infection, and obesity. Patient 3 also had high fever and cough at the onset of the illness. She had visited a chicken market 1 week before the onset of symptoms. The demographic and epidemiologic

characteristics of the three patients are summarized in Table 1TABLE 1

Demographic,

Epidemiologic, and Virologic Characteristics and Complications, Treatment, and Clinical Outcomes of Three Patients Infected with Avian-Origin Influenza A (H7N9) Virus..

Determination of Causative Pathogens We confirmed, by means of real-time RT-PCR, viral isolation, and full genome sequencing, that all three patients were infected with a novel avian-origin influenza A (H7N9) virus. Original clinical samples obtained from all three patients were confirmed, by means of real-time RT-PCR, to be positive for H7N9 and negative for seasonal influenza viruses (H1, H3 or B), H5N1, SARS-CoV, and HCoV-Erasmus Medical Center (EMC). Influenza viruses A/Shanghai/1/2013 (H7N9), A/Shanghai/2/2013 (H7N9), and A/Anhui/1/2013 (H7N9) were isolated from Patients 1, 2, and 3, respectively. Complete sequences of the three H7N9 influenza viruses showed that they were 97.7 to 100% identical in all eight gene segments (see Table S1 in theSupplementary Appendix). Phylogenetic analysis of all genes of the isolates showed that each gene

was of avian origin (Figure 1FIGURE 1

Phylogenetic Trees of Genes of H7N9 Influenza A

Viruses., and Figure S1 in theSupplementary Appendix). The gene encoding hemagglutinin (HA) shared

the highest identity with A/duck/Zhejiang/12/2011 (H7N3, subtype ZJ12). The gene encoding neuraminidase (NA) protein was most closely related to A/wild bird/Korea/A14/2011 (H7N9, subtype KO14); however, the HA gene from the H7N9 viruses in our three patients was highly divergent from that in the KO14 virus. All six internal genes shared the highest similarity with A/brambling/Beijing/16/2012-like viruses (H9N2) (Figure 1). Phylogenetic results indicated that it was a triple reassortant H7N9 virus (Figure

2FIGURE 2

Hypothetical Host and Lineage Origins of the Gene Segments of the Novel

Reassortant Human Influenza A (H7N9) Viruses.).

In all three viruses, the HA cleavage site possesses only a single amino acid R (arginine), indicating low pathogenic effects in poultry. A T160A mutation was identified at the 150-loop (H3 numbering) in the HA gene of all three viruses. Substitution Q226L at the 210-loop in the HA gene was found in both the A/Anhui/1/2013 and A/Shanghai/2/2013 viruses but not in the A/Shanghai/1/2013 virus (Table 2TABLE 2

Molecular Analysis of Three of the 2013 H7N9 Viruses.). Five amino acids were deleted in

the stalk region of NA residue 69 to 73. The M2 protein contained the S31N substitution, indicating resistance to amantadine. Other mutations 89V and E627K in PB2 and 42S in NS1 were also identified (Table 2). The amino acids in A/Shanghai/1/2013, which differed from those in A/Anhui/1/2013 and A/Shanghai/2/2013, are shown in Table S2 in theSupplementary Appendix. To date, five additional H7N9 viruses have been isolated from five patients. Sequencing analysis indicates that all five viruses are highly similar to both A/Shanghai/2/2013 and A/Anhui/1/2013. Some variability is observed, such as Q226L in HA and R292K in NA. On the basis of these data, diagnostic tests for the novel reassortant H7N9 viruses have been developed. The specific sequences are available on the website of the World Health Organization (www.who.int/influenza/gisrs_laboratory/a_h7n9/en/). Clinical Features and Outcomes of the Patients The clinical characteristics of the patients are shown in Table S3 in theSupplementary Appendix. Fever and cough were the most common symptoms. The white-cell count was normal or slightly decreased. Elevated levels of aspartate aminotransferase, creatine kinase, and lactate dehydrogenase were observed in all the patients. Bilateral ground-glass opacities and consolidation were detected on chest radiography

(Figure 3FIGURE 3

Chest Radiographs.).

Several complications of the illness were observed. All the patients had ARDS. Patient 3 had septic shock and acute renal damage. Carbapenem-resistant Acinetobacter baumannii was cultured from lower respiratory tract specimens obtained from two of the patients after the initiation of mechanical ventilation. Combination antibiotic therapy, glucocorticoids, and intravenous immunoglobulin were administered in all three patients. Antiviral therapy was initiated 6 to 7 days after the onset of illness (Table 1). Patient 1 declined admission to the intensive care unit (ICU) and intubation. He died from refractory hypoxemia 13 days after the onset of illness. Patient 2 was admitted to the ICU and intubated 48 hours after

admission owing to progressive dyspnea. He died from refractory hypoxemia after 4 days in the ICU. ARDS and septic shock developed in Patient 3 on day 6 after the onset of illness. She was admitted to the ICU, and extracorporeal membrane oxygenation was initiated. She died on April 9.

DISCUSSION
We have identified a novel reassortant influenza A (H7N9) virus that is associated with severe human infection. Currently, only 25 H7N9 viruses are available in GenBank. The H7N9 viruses we identified in the three patients were of avian origin, but only the NA gene was closely related to that from another H7N9 virus (KO14). The HA gene was similar to that of an H7N3 virus (ZJ12) from a nearby region (Zhejiang Province) in China. All the internal gene segments were closely related to those from avian H9N2 viruses, particularly a virus isolated from a brambling in Beijing (BJ16) (Figure 1, and Figure S1 in the Supplementary Appendix). Thus, the human H7N9 viruses are the product of reassortment of viruses that are of avian-origin only. In addition, the phylogenetic trees showed that A/Shanghai/1/2013 is phylogenetically distinct from A/Anhui/1/2013 and A/Shanghai/2/2013 across all gene segments, which suggests that there have been at least two introductions into humans (Figure 1, and Figure S1 in the Supplementary Appendix). Currently, there are no data to suggest that this reassortment occurred in a mammalian host, and the similarity of the human viruses to avian viruses may be stronger support for direct avian transmission of this virus. However, influenza surveillance of birds, swine, and humans is limited in China and nearby countries, making it difficult to resolve this question. Although human infections with avian-origin H7 avian influenza viruses have been observed before,1,2,5,10,11 infection of humans with an N9 subtype influenza virus has not been reported previously. Human H7 influenza infections are generally mild, causing conjunctivitis or modest respiratory symptoms, although a fatal case was reported before this H7N9 outbreak.5 All three cases of H7N9 infection reported here were virulent, with the patients' conditions deteriorating rapidly with the development of severe pneumonia and ARDS, and ultimately resulted in death. All the patients had preexisting medical conditions, and two had a history of direct contact with poultry. Two patients presented with rhabdomyolysis, which has rarely been reported in patients infected with H1N1 or H5N112 influenza viruses. Encephalopathy, which is normally more common in pediatric patients with influenza,13 was observed in two patients. The affinity of the influenza virus to different sialyl-sugar structures is an important determinant of range and pathogenicity in the viral host.14,15 Human influenza viruses preferentially bind to 2,6 sialyl glycan, whereas most avian viruses bind to 2,3 sialyl glycan.16,17 Q226L in the HA protein, which was first reported in H7 field viruses, as well as H5 subtypes, was expected to bind strongly to -2,6 human-like receptors. A laboratory-produced Q226L mutation at the 210-loop of HA has been shown to change the receptor binding of avian origin to a human-type receptor binding and might increase the ability of the virus to be transmitted by air, as reported previously.18,19 Moreover, the lack of a glycosylation site on the 150loop might decrease the affinity to -2,3 avian-like receptors. The effects of these mutations require further study. A deletion of five amino acids in the viral NA stalk has been observed in the novel reassortant H7N9 viruses. A similar deletion in the H5N1 avian virus has been shown to be responsible for the change in viral tropism to the respiratory tract20 or to enhance viral replication,21 and it has been suggested that this deletion may be associated with adaptation and transmission in domestic poultry. 22,23Since April 4, it has been reported that H7N9 viruses similar to those isolated from the three patients described here have been

isolated from pigeons and chickens, indicating that the novel H7N9 viruses might currently be circulating in poultry. Moreover, the E627K substitution in the PB2 gene has been associated with increased virulence in mice and was reported to be associated with improved replication of avian influenza viruses in mammals.24,25 A combination of these substitutions may contribute to the human infection and severe disease. Other possible virulence molecular markers are shown in Table 2. The potential virulence mutations are described on the basis of previous studies in animals, but the pathogenesis in humans remains unknown. The difference between the two Shanghai viruses and the similarity between the Shanghai/2 and Anhui/1 viruses argue against human-to-human transmission in these cases, and no close contacts of the patients have tested positive for these viruses. However, limited human-to-human transmission was observed in the H7 outbreak in the Netherlands in 200310; therefore, the pandemic potential of these novel avian-origin viruses should not be underestimated. Currently there is no vaccine available for these novel viruses, and it is not known whether the current candidate H7 vaccine viruses, of which three are North American viruses and the other three are avian viruses from 2000 in the Netherlands, may be effective. The influenza H7N9 A/Anhui/1/2013 strain has been proposed to be one of the candidate vaccine strains since it grows to a very high titer in eggs. Heightened protective measures should be taken when dealing with these viruses, and increased surveillance and analyses of these viruses are needed. Severe avian influenza A (H7N9) infections, characterized by high fever and severe respiratory symptoms, may pose a serious human health risk. We are concerned by the sudden emergence of these infections and the potential threat to the human population. An understanding of the source and mode of transmission of these infections, further surveillance, and appropriate counter measures are urgently required.
Supported

he first identified cases of human infection with a novel influenza A (H7N9) virus occurred in eastern China during February and March 2013 and were characterized by rapidly progressive pneumonia, respiratory failure, acute respiratory distress syndrome (ARDS), and fatal outcomes.1We analyzed available data from field investigations to characterize the descriptive epidemiology of laboratory-confirmed cases of avian influenza A (H7N9) virus infection in humans reported to the Chinese Center for Disease Control and Prevention (China CDC) as of April 17, 2013. In this report, we summarize the preliminary findings of case investigations and follow-up monitoring of close contacts of persons with confirmed cases of H7N9 virus infection who have been identified to date. This is an ongoing investigation.

METHODS
Case Definitions The case definitions of suspected and confirmed human infection with H7N9 virus were based on the H5N1 case definitions, as recommended by the World Health Organization (WHO) in 2006 (Section S1 in the Supplementary Appendix, available with the full text of this article at NEJM.org).2 The laboratory test assays for H7N9 virus that we performed have been described previously (Section S2 in the Supplementary Appendix).3 Identification of Cases

Suspected cases of H7N9 virus infection were identified through the Chinese surveillance system for pneumonia of unexplained origin, which was established in 2004.4 Beginning on April 3, 2013, enhanced surveillance was implemented for suspected cases of H7N9 virus infection among persons with mild or moderate illness.5 Persons with suspected cases of H7N9 virus infection with mild or moderate illness were identified from the Chinese sentinel surveillance system for influenza-like illness, which has been described previously.6 Once each suspected case of H7N9 virus infection was identified, the local CDCs, including prefecture and provincial CDCs, conducted the initial field investigations and obtained respiratory specimens, which were shipped to the National Influenza Center of the China CDC in Beijing for H7N9 laboratory testing. A field investigation team comprising staff members of the China CDC and or local CDC conducted field investigations of the confirmed cases of H7N9 virus infection. Data Collection It was determined by the National Health and Family Planning Commission that the collection of data from H7N9 case patients and their close contacts was part of a continuing public health investigation of an outbreak and was exempt from institutional review board assessment. Data were collected through a review of medical records and interviews with relatives, contacts, and health care workers who provided medical care for the case patients. We collected information on the dates of illness onset, visits to clinical facilities, hospitalization, and clinical outcomes. Epidemiologic data were collected through interviews and field observations and were reported to the China CDC. Investigators interviewed the relatives of each patient with a confirmed case of H7N9 virus infection to determine exposure histories during the 2 weeks before the onset of the illness, including the dates, times, frequency, and patterns of exposures to poultry or other animals such as swine and wild birds. All epidemiologic information that was collected during the field investigations, including exposure history, timelines of events, and identification of close contacts, was cross-validated, since we were unable to interview any critically ill H7N9 case patients. Households and places that were known to have been visited by the case patients in the 2 weeks before the onset of illness were investigated to assess exposures to poultry and swine, as well as environmental exposures. Identification and Follow-up of Close Contacts We defined close contacts of patients with confirmed H7N9 virus infection as described previously for H5N1 field investigations7 (Section S3 in the Supplementary Appendix); once we identified the close contacts, we monitored them daily for 7 days for symptoms of illness and collected throat swabs from contacts in whom symptoms developed to test for the presence of the H7N9 virus. Antiviral chemoprophylaxis was not provided to close contacts. Paired serum samples were obtained from patients with suspected H7N9 virus infection who did not have respiratory specimens available for H7N9 serologic testing. Oseltamivir treatment was recommended for close contacts in whom symptoms developed (Section 3 in the Supplementary

Appendix). Data on demographic characteristics and exposure were collected for close contacts. Statistical Analysis
We used descriptive statistics to summarize the epidemiologic characteristics and H7N9 testing results for persons with suspected cases of H7N9 virus infection, for those with confirmed cases, and for close contacts of those with confirmed cases. The methods we used for estimating the incubation period have been described previously.8

RESULTS
Epidemiologic Characteristics of Confirmed Cases

From March 25 through April 17, 2013, respiratory specimens from 664 hospitalized patients with pneumonia of unexplained origin were tested, and 81 patients (12.2%) were confirmed to be infected with the H7N9 virus. Of 5551 respiratory specimens obtained from outpatients with an influenza-like illness through the sentinel surveillance system for influenza-like illness, 1 (0.02%) tested positive for the H7N9 virus. As of April 17, 2013, a total of 82 laboratory-confirmed cases of H7N9 virus infection and 2 suspected

cases had been identified (Figure 1FIGURE 1

Date of Onset of Illness in First 82 Patients

with Confirmed H7N9 Virus Infection, According to Province in China. and Figure 2FIGURE 2

Geographic Distribution of 82 Confirmed and 2 Suspected Cases of H7N9 Virus Infection in China, as of April 17, 2013.); cases were identified in the following provinces: Shanghai (31 confirmed cases

and 1 suspected case), Zhejiang (25 confirmed cases), Jiangsu (20 confirmed cases and 1 suspected case), Anhui (3 confirmed cases), Henan (2 confirmed cases) and Beijing (1 confirmed case). The median age of patients with confirmed H7N9 virus infection was 63 years (range, 2 to 89); 38 cases (46%) occurred in persons 65 years of age or older, and 2 (2%) were in children younger than 5 years of age, both of whom had clinically mild upper respiratory illness (see Figure S1 in the Supplementary Appendixfor the age distribution). Most confirmed cases occurred in males (73%), 84% of the case patients were urban residents, and 54 of 71 patients with available data (76%) had underlying medical conditions (Table

1TABLE 1

Epidemiologic Characteristics of 82 Patients with Confirmed H7N9 Virus Infection in

China.). Among 46 of 54 case patients with sufficient data for a more specific classification of underlying

conditions, 40 (87%) were considered to be at increased risk for influenza complications owing to age (<5 years or 65 years) or prevalence of certain underlying medical conditions.9 Four of the patients with confirmed cases (5%) worked as poultry workers: 3 slaughtered poultry at a live poultry market, and 1 transported live poultry. A total of 81 of 82 patients with confirmed H7N9 virus infection (99%) were hospitalized. Among the 51 patients with confirmed cases for whom data were available, isolation precautions were instituted for 33 (65%) in an intensive care unit (ICU) because of severe lower respiratory tract disease. As of April 17, a total of 17 patients with confirmed H7N9 virus infection (21%) and 1 patient with suspected infection had died of acute respiratory distress syndrome (ARDS) or multiorgan failure, and 60 patients with confirmed cases and 1 with a suspected case remained critically ill; 4 with clinically mild cases had been discharged from the hospital, and 1 pediatric patient had not been admitted to the hospital. Among 82 confirmed cases of H7N9 virus infection, 7 (9%) were confirmed by means of virus isolation, 2 (2%) by means of serologic testing, and 73 (89%) by means of nucleic acid detection. Viral culture of 73 respiratory specimens that were confirmed as positive by means of real-time reverse-transcriptasepolymerase-chain-reaction (RTPCR) assays and diagnostic testing of specimens from suspected cases are ongoing.

Data on recent exposure to animals were available for 77 of the 82 patients with confirmed H7N9 virus infection. Of these, 59 (77%) reported a history of recent exposure to animals, (Table 1): 45 (76%) to chickens, 12 (20%) to ducks, and 4 (7%) to swine; the exposures occurred either while they were working at or visiting a live animal market. Other animals that these 59 patients reported having been exposed to included pigeons, geese, quail, wild birds, pet birds, cats, and dogs. Information on a history of exposure to live animals is unclear for 5 patients with confirmed H7N9 virus infection, since the investigations are still ongoing. The estimated median incubation period in 23 patients with confirmed cases for whom detailed data on animal and environmental exposures was available was 6 days (range, 1 to 10) (Table S1 in the Supplementary Appendix). Family Clusters As of April 17, three family clusters had been identified in two provinces. Detailed exposure and timeline information are available for two family clusters that were identified early (Figure 3FIGURE 3

Timeline of Pertinent Exposures and Dates of Illness Onset in Two Family Clusters of Cases of H7N9 Virus Infection in China., and Section S4 in the Supplementary Appendix); the investigation of one

cluster in Shanghai is still ongoing. The first family cluster comprised two persons with confirmed H7N9 virus infection and one with a suspected infection. The index case patient and his father (confirmed case 2) lived in the same house, and the index patient's brother (confirmed case 1) lived with his wife nearby. After the index case patient (suspected case 1) became ill, his brother (confirmed case 1) and his father (confirmed case 2) had prolonged, close, unprotected contact with him, including eating together, providing care, and accompanying him to seek medical care before his hospitalization (Section S4 in the Supplementary Appendix). None of the three members of the cluster raised poultry or other animals, none brought live poultry into their home, and none had direct contact with sick or dead poultry. The index case patient had visited a live poultry market, purchased a chicken, observed the slaughtering process, brought the freshly killed chicken home, and prepared, cooked, and ate the chicken within 2 weeks before the onset of his illness. Another family cluster was identified that included one person with confirmed H7N9 virus infection and one person with a suspected case (Figure 2, and Section S4 in the Supplementary Appendix). After the father (suspected case) became ill, his daughter (confirmed case) had prolonged, close, unprotected contact with him, including eating together, providing care, and accompanying him to seek medical care before his hospital admission. She also provided unprotected bedside hospital care for her father during the period from March 11 through March 15. Diarrhea developed in the father, and the daughter washed her father's diarrhea-soiled underwear on March 18 while wearing gloves. The father had visited a live poultry market 7 days before the onset of his illness. The daughter did not raise poultry or animals at home and had not had any exposures to animals (i.e., had not brought live poultry into the home or visited a live poultry market or had any direct or indirect contact with poultry or pigs). Medical Care Timelines

Among the 81 patients with confirmed H7N9 virus infection for whom data were available (99% of the 82 patients with confirmed cases), the median time from the onset of illness to the first medical visit was 1 day,

and patients were hospitalized a median of 4.5 days after the onset of illness (Table 2TABLE 2
Clinical Characteristics and Medical Care Timelines for 82 Patients with Confirmed H7N9 Virus Infection in China.). Among 64 of the patients with available data on oseltamivir administration, 41 (64%) received

oseltamivir treatment beginning a median of 6 days after the onset of illness. ARDS developed during the course of hospitalization in 19 of 40 patients with confirmed H7N9 virus infection for whom data were available (48%) after a median of 8 days, and 17 patients died a median of 11 days after the onset of illness. Close Contacts As of April 17, data were available for 1689 close contacts of the 82 patients with confirmed cases in Shanghai (435 contacts), Jiangsu (448), Zhejiang (676), Anhui (100), Henan (28), and Beijing (2). Among the 678 close contacts of 33 patients with confirmed cases for whom demographic information was available, 422 (62%) were health care workers, 134 (20%) were family member or relatives, and 122 (18%) were social contacts. As of April 17, a total of 1251 of the 1689 contacts had been followed for up for 7 days; among these close contacts (not including those in the family clusters), respiratory symptoms developed during the 7-day surveillance period in 19 (1.5%): 2 household members, 1 medical intern, 1 patient who shared a room with a confirmed case patient, and 15 health care workers (Section S5 in the Supplementary Appendix). All the throat swabs collected from these 19 ill contacts a median of 1 day (range, 0 to 7) after the onset of illness were negative for the H7N9 virus, as assessed by means of realtime RT-PCR.

DISCUSSION
An epidemiologic study of 82 confirmed cases of H7N9 virus infection in China among persons with illness onset during the period from February through April 17, 2013, indicates that the infection affected persons in a wide age spectrum and caused severe lower respiratory tract illness. To date, the mortality is 21%, but since many of patients with confirmed H7N9 virus infection remain critically ill, we suspect that the mortality may increase. Except for one family cluster with 2 confirmed cases, patients with confirmed H7N9 virus infection were epidemiologically unrelated and were identified in six areas of China. Most of the patients with confirmed H7N9 virus infection were considered to be at increased risk for complications from influenza owing to age (<5 years or 65 years; median age of patients with confirmed cases, 63 years) or the prevalence of certain underlying medical conditions. Human infections with influenza A (H7) viruses have been reported sporadically and are usually associated with exposures to poultry.10-12 Previous human cases of H7 virus infection have been characterized by mild illness (conjunctivitis or uncomplicated influenza) or moderate illness (lower respiratory tract disease) that results in hospitalization.10,11,13,14 Only one fatal case of H7 virus infection has been reported previously; that case occurred in an adult with a highly pathogenic avian influenza A (H7N7) virus infection.15 Many of the confirmed H7N9 case patients had critical and fatal illness, suggesting that the

H7N9 virus is more virulent in humans than are other H7 viruses. The H7N9 case fatality proportion to date is lower than that for reported cases of H5N1 virus infection.16However, early surveillance for H7N9 cases was focused on case finding for severe lower respiratory tract illness, and since April 3, expanded testing of outpatients with influenza-like illness has identified some mild cases of illness with H7N9 virus infection. Enhanced surveillance for less severe illness with H7N9 virus infection will help to determine the clinical spectrum of the illness and the total number of cases of H7N9 symptomatic illness and to inform an understanding of the true case fatality proportion. Since this H7N9 virus appears to have emerged recently to infect humans, population immunity is expected to be low, and persons of any age may be susceptible to infection. Although the source of the H7N9 virus infection in patients with confirmed cases who had exposure to animals cannot be verified without extensive H7N9 testing of animals, we suspect that it is likely to be infected poultry; additional studies are needed. No animal outbreaks were identified in the areas with confirmed H7N9 cases, but 77% of cases with available data occurred in patients who had exposure to live animals such as poultry or swine, including during visits to live animal markets. This raises the possibility of zoonotic H7N9 virus transmission from healthy-appearing swine or poultry to humans through direct or close contact or through exposure to environments that are contaminated with infected swine or poultry. For example, visiting a live poultry market, where avian influenza A viruses can be maintained and amplified, has been identified as a risk factor for H5N1 virus infection in Hong Kong 17 and urban China.18,19 However, casecontrol studies are needed to identify risk factors for H7N9 virus infection. Until the source of H7N9 virus infection is known, implementation of control measures at live poultry markets, such as a ban on the selling of live poultry in market stalls or even market closure, poultry culling, and market disinfection measures that have been taken to control the spread of H5N1 virus may be considered in order to help control potential zoonotic transmission of H7N9 virus. Follow-up prospective investigations of close contacts of patients with confirmed H7N9 virus infection have not conclusively established human-to-human H7N9 transmission of the virus from one confirmed case to another to date. However, in two family clusters, limited human-to-human transmission of H7N9 virus after close, prolonged, unprotected contact with a symptomatic patient with suspected H7N9 virus infection remains a possibility, because specimens were not available for H7N9 testing from patients with suspected cases; one patient with a suspected case died before a specimen could be obtained, and H7N9 testing of the other patient with a suspected case is still ongoing. Similar family clusters of H5N1 cases that occurred after common poultry exposures or limited human-to-human transmission have been identified.7,20,21 Paired serum samples are being obtained during the acute and convalescent stages of illness from contacts of case patients for further assessment of the potential for secondary human-to-human H7N9 virus transmission, including the identification of asymptomatic infections. Although the risk of humanto-human transmission of H7N9 virus appears to be low, the actual risk is currently unknown, and the Chinese national guidelines recommend implementing control measures, such as prompt isolation of the patient, active monitoring of close contacts, and implementation of standard, contact, and droplet precautions by health care personnel in hospitals. In addition, national guidelines recommend that antiviral treatment with oseltamivir should be administered as soon as possible in patients with suspected or confirmed cases of H7N9 virus infection.

The median time from the onset of illness to hospitalization among the 81 of 82 patients with confirmed H7N9 virus infection for whom data on hospitalization were available was 4.5 days, and the median time from the onset of illness to the development of ARDS among the 19 case patients with ARDS (out of 40 patients for whom data on ARDS were available) was 8 days; the corresponding median times among patients with H5N1 virus infection were 7 days and 7.5 days.22The median duration from the onset of illness to death among the 17 persons with confirmed cases who died was 11 days. The initial findings suggest that H7N9 virus infection can cause critical illness and fatal disease and may affect persons in a wider age range than the H5N1 virus has in China to date (Figure S1 in the Supplementary Appendix). Patients with confirmed cases received oseltamivir antiviral treatment a median of 6 days after the onset of illness (median before April 3, 9 days), probably owing to delayed suspicion of influenza. Retrospective observational studies of influenza A (H1N1)pdm09 and H5N1 virus infections suggest that early oseltamivir treatment probably has the greatest clinical benefit but that starting treatment up to 5 days after the onset of illness may still reduce the risk of critical illness and death.23-27 Preliminary data suggest that the H7N9 viruses isolated from humans and analyzed to date are resistant to adamantane antiviral agents and are susceptible to neuraminidase inhibitors. Early clinical suspicion of H7N9 virus infection and early administration of oseltamivir may help to reduce the severity of the disease. Our study had several limitations. First, we did not collect detailed information from all patients on exposures, such as the times, frequency, intensity, and duration of exposures. Information on exposures is useful for estimating the incubation period after possible exposure to animals or live-animal markets and for evaluating the risk factors for H7N9 virus infection. Second, we may not have identified all the close contacts of case patients and were not able to conduct active follow-up of all contacts. As of April 17, 2013, complete follow-up data were not available for some of the close contacts. Third, we did not have a standard protocol and questionnaire to collect information from all contacts of the first 82 patients with confirmed cases. However, the China CDC has issued a guideline and protocol for field investigations of case patients and close contacts and since April 1 has provided training for personnel at all 31 provincial CDCs. This will help ensure standard data collection. Fourth, specimens were not available for H7N9 testing from some patients with suspected cases. Clinical outcomes in the 82 patients with confirmed H7N9 virus infection are reported as of April 17, 2013, and 60 case patients remain hospitalized. Paired serum samples have not been obtained from some of the contacts; no serologic testing results are available at this time, and given the fact that it is early in the investigation, more time is needed to allow for a humoral immune response in serum obtained during the convalescent period and to allow time for serologic testing to be performed. In summary, a novel influenza A (H7N9) virus has caused severe and fatal illness in persons in six different areas of China to date. Some clinically mild cases have been identified since the surveillance was widened, suggesting that there is a wide clinical spectrum of H7N9 virus infection. The initial epidemiologic findings suggest that most confirmed H7N9 cases were epidemiologically unrelated. Follow-up investigations of contacts of patients with confirmed H7N9 virus infection suggest that the risk of secondary H7N9 virus transmission, including to health care personnel, is low at this time. However, in two family clusters that include persons with confirmed H7N9 virus infection and persons with epidemiologically linked suspected cases, limited nonsustained human-to-human H7N9 virus transmission could not be ruled out and may have occurred among blood-related family members. Enhanced surveillance for severe and mild human

illness with H7N9 virus infection is needed to determine the clinical spectrum of the infection and the total number of symptomatic H7N9 infections. Casecontrol studies to identify risk factors and continued investigations of case patients and their contacts are indicated. Data from investigations of potential animal and environmental sources are urgently needed to inform public health control measures.

BACKGROUND
Both genetic variation at the 17q21 locus and virus-induced respiratory wheezing illnesses are associated with the development of asthma. Our aim was to determine the effects of these two factors on the risk of asthma in the Childhood Origins of Asthma (COAST) and the Copenhagen Prospective Study on Asthma in Childhood (COPSAC) birth cohorts.

METHODS
We tested genotypes at the 17q21 locus for associations with asthma and with human rhinovirus (HRV) and respiratory syncytial virus (RSV) wheezing illnesses and tested for interactions between 17q21 genotypes and HRV and RSV wheezing illnesses with respect to the risk of asthma. Finally, we examined genotypespecific expression of 17q21 genes in unstimulated and HRV-stimulated peripheral-blood mononuclear cells (PBMCs).

RESULTS
The 17q21 variants were associated with HRV wheezing illnesses in early life, but not with RSV wheezing illnesses. The associations of 17q21 variants with asthma were restricted to children who had had HRV wheezing illnesses, resulting in a significant interaction effect with respect to the risk of asthma. Moreover, the expression levels ofORMDL3 and of GSDMB were significantly increased in HRV-stimulated PBMCs, as compared with unstimulated PBMCs. The expression of these genes was associated with 17q21 variants in both conditions, although the increase with exposure to HRV was not genotype-specific.

CONCLUSIONS
Variants at the 17q21 locus were associated with asthma in children who had had HRV wheezing illnesses and with expression of two genes at this locus. The expression levels of both genes increased in response to HRV stimulation, although the relative increase was not associated with the 17q21 genotypes. (Funded by the National Institutes of Health.)

BACKGROUND
Critical illness is often accompanied by hypercortisolemia, which has been attributed to stress-induced activation of the hypothalamicpituitaryadrenal axis. However, low corticotropin levels have also been reported in critically ill patients, which may be due to reduced cortisol metabolism.

METHODS
In a total of 158 patients in the intensive care unit and 64 matched controls, we tested five aspects of cortisol metabolism: daily levels of corticotropin and cortisol; plasma cortisol clearance, metabolism, and production during infusion of deuterium-labeled steroid hormones as tracers; plasma clearance of 100 mg

of hydrocortisone; levels of urinary cortisol metabolites; and levels of messenger RNA and protein in liver and adipose tissue, to assess major cortisol-metabolizing enzymes.

RESULTS
Total and free circulating cortisol levels were consistently higher in the patients than in controls, whereas corticotropin levels were lower (P<0.001 for both comparisons). Cortisol production was 83% higher in the patients (P=0.02). There was a reduction of more than 50% in cortisol clearance during tracer infusion and after the administration of 100 mg of hydrocortisone in the patients (P0.03 for both comparisons). All these factors accounted for an increase by a factor of 3.5 in plasma cortisol levels in the patients, as compared with controls (P<0.001). Impaired cortisol clearance also correlated with a lower cortisol response to corticotropin stimulation. Reduced cortisol metabolism was associated with reduced inactivation of cortisol in the liver and kidney, as suggested by urinary steroid ratios, tracer kinetics, and assessment of liverbiopsy samples (P0.004 for all comparisons).

CONCLUSIONS
During critical illness, reduced cortisol breakdown, related to suppressed expression and activity of cortisolmetabolizing enzymes, contributed to hypercortisolemia and hence corticotropin suppression. The diagnostic and therapeutic implications for critically ill patients are unknown. (Funded by the Belgian Fund for Scientific Research and others; ClinicalTrials.gov numbers, NCT00512122 andNCT00115479; and Current Controlled Trials numbers,

A 59-year-old man presented with a 6-week history of pain in the thumb. He also reported having a cough, weight loss, and a history of heavy smoking. Physical examination revealed swelling, redness, and tenderness over the first right metacarpal and a mass (10 cm in diameter) fixed to the thorax. Laboratory tests revealed a C-reactive protein level of 123 mg per liter and a leukocyte count of 34109 per liter. A radiograph of the hand showed osteolysis of the first metacarpal (Panel A, arrow), and a chest radiograph showed a solid mass of the thoracic wall expanding into the subpleural space (Panel B, long arrow) and a tumor in the hilus of the right lung (Panel B, short arrow). A specimen of the metacarpal lesion obtained through needle aspiration was negative for bacteria. The final diagnosis was lung adenocarcinoma with osseous lesions of the hand and ribs. The patient received palliative care.

In a recent study that analyzed data from the GeoSentinel Surveillance Network (which consists of 42 specialized travel or tropical-medicine sites located around the world) on 25,867 returned travelers over a 9-year period (from 1996 to 2005), of the 2902 clinically significant pathogens that were isolated, approximately 65% were parasitic, 31% bacterial, and 3% viral. Six organisms (giardia, campylobacter, Entamoeba histolytica, shigella, strongyloides, and salmonella species) accounted for 70% of the gastrointestinal burden.

Clinical Pearls

- What are the clinical manifestations of Giardia lamblia infection? Giardia lamblia (also called Giardia intestinalis or Giardia duodenalis) is highly contagious (ingestion of as few as 10 to 25 cysts may cause disease), with persons becoming infected through the ingestion of cysts in contaminated food or water. However, person-to-person transmission is possible. The clinical manifestations range from mild intestinal problems that resolve spontaneously to complex symptoms that last up to several weeks, such as protein-losing enteropathy, postinfectious fatigue, chronic diarrhea, abdominal pain, nausea, and weight loss. In children, the disease can cause growth and cognitive impairment as a result of iron and micronutrient deficiencies. - What is the natural history, typical clinical course, and methods to diagnose amebiasis? E. histolytica and E. moshkovskii are pathogenic in humans, causing amebiasis. The parasite is acquired through the ingestion of food or water contaminated with fecal cysts. After it has been ingested, the cyst emerges in the terminal ileum as an active trophozoite, which migrates to the colon where it colonizes the mucus layer. Invasion may take days to years after the initial infection and is characterized by fever, abdominal pain, and bloody dark-brown diarrhea. However, 90% of cases are asymptomatic and self-limiting. Symptomatic disease occurs when trophozoites invade the mucosa and submucosa, and some trophozoites enter the portal circulation and disperse to the liver and other soft organs. Disease of the right colon is common and is associated with the following serious complications: strictures, rectovaginal fistulas, bowel obstruction, toxic megacolon, perforation, peritonitis, and death. Only 1% of clinical cases of amebiasis involve the liver. Several stool antigen assays specific for E. histolytica are commercially available to make an accurate diagnosis of intestinal or hepatic amebiasis on the basis of the Gal/GalNAc lectin. Microscopic examination of the stool is no longer performed for amebiasis because of its low sensitivity and specificity; with microscopy, it is easy to confuse E. histolytica with the identically appearing and much more common nonpathogenic parasite E. dispar.

Morning Report Questions Q: What are the manifestations of strongyloides infection? A: Strongyloides stercoralis (threadworm) is the most dominant species causing infection in humans. Third-stage filariform larvae penetrate the skin (usually the foot) of the human host, reach the lungs via the blood circulation, and enter respiratory pathways. From there, they migrate upward through the trachea, are swallowed, and finally reach the small intestine, where they mature into adult egg-laying female worms. Female worms embed in the submucosa of the duodenum,

where they produce dozens of eggs per day. These hatch in the gut lumen, and the first-stage rhabditiform larvae either are passed out in the feces and develop into infective third-stage larvae or remain in the gastrointestinal tract of the human host and start a new infection cycle (autoinfection). Autoinfection can result in persistent infection for decades. More than 50% of patients with a chronic infection are asymptomatic. For a subset of patients with disease, the symptoms include erythematous pruritus, skin eruptions, larva currens, abdominal pain, diarrhea, and weight loss. In travelers presenting with eosinophilia or elevated IgE levels, strongyloides should be considered in the differential diagnosis. In immunocompromised persons, strongyloidiasis can cause a hyperinfection syndrome owing to the reproductive capacity of the parasite inside the host. In cases of disseminated disease, the hyperinfection syndrome can be associated with a mortality rate of close to 90%. Q: What is the natural history and clinical presentation of schistosomiasis? A: Schistosomiasis is a common chronic helminth disease caused by intravascular parasitic schistosoma trematode worms. The three most important species in humans are Schistosoma hematobium, S. mansoni, and S. japonicum. Schistosome transmission requires the contamination of water by egg-containing feces or urine, a specific freshwater snail as intermediate host, and human contact with water inhabited by the intermediate host snails. Schistosome larvae (cercariae) emerge from the snails and penetrate human skin, thereby instigating infection. A maculopapular eruption consisting of discrete erythematous, raised lesions that vary in size from 1 to 3 cm may arise at the site of percutaneous penetration by the cercariae. Patients with acute schistosomiasis, or Katayama fever, which usually begins with the deposition of schistosome eggs into host tissues, can present with fever, malaise, myalgia, fatigue, nonproductive cough, diarrhea (with or without blood), hematuria (S. hematobium), and right-upper-quadrant pain. A skin reaction may develop within a few hours after infection in migrants or tourists infected for the first time, although a rash may appear as much as a week later. In cases of infection with S. mansoni and S. japonicum, a T-cell-mediated granulomatous reaction to schistosome eggs leads to fibrosis and chronic disease of the human liver, resulting in the development of severe hepatosplenic schistosomiasis; in cases of S. hematobium, this reaction leads to fibrosis and calcification of the bladder and ureters, which can result in bladder cancer.

TEACHING TOPIC Vaginal Bleeding CASE RECORDS OF THE MASSACHUSETTS GENERAL HOSPITAL, Case 14-2013: A 70-Year-Old Woman with Vaginal Bleeding, (https://www.nejm.org/action/doSecureKeyLogin?uuid=7676846&dateTime=2013051 80000&key=0rSCw5hWJYFuQ6MwpDd%2FHHb9duCvOf9ptOHi%2FlfDIO0%3D&uri=/doi/full/

10.1056/NEJMcpc1209276?query=BUL) R.T. Penson and Others CME Exam

The underlying cause of abnormal vaginal bleeding is age-dependent. Ten percent of premenopausal women with abnormal bleeding have a malignant tumor. In contrast, 75% of women over 70 years of age with postmenopausal bleeding have cancer, and the risk rises with age in postmenopausal women.

Clinical Pearls - What is the typical presentation of carcinosarcoma of the uterus? Postmenopausal vaginal bleeding is the most common manifestation of carcinosarcoma. Patients with carcinosarcoma also frequently present with the classic triad of painful postmenopausal bleeding, an enlarged uterus, and prolapsed tumor visible at the cervical os. - Under what circumstances is surgery not the primary treatment for uterine cancer? In only a few circumstances is surgery not the primary treatment for uterine cancer -- when there is a desire to preserve fertility, high operative risk, and unresectable disease. The goals of surgical treatment are excision of all disease with at least a 1-cm margin and staging of the tumor. The initial spread is to regional lymph nodes; therefore, standard treatment is a radical total abdominal hysterectomy and bilateral salpingo-oophorectomy with lymphadenectomy. Endometrial cancers have several potential patterns of spread: direct invasion and expansion of the primary tumor, lymphatic invasion, hematogenous spread, and intraperitoneal dissemination. Because metastasis is common, preoperative combination positron-emission tomography and computed tomography (PET-CT) and a meticulous exploratory laparotomy are standard practice.

Morning Report Questions Q: What features affect the overall prognosis of patients with carcinosarcoma? A: Diagnostic features of malignant mixed mullerian tumor (carcinosarcoma) include the finding of a biphasic malignant tumor that is composed of high-grade carcinoma (most commonly endometrioid or serous) and sarcoma and is typically homologous (arising from mesenchymal tissue normally found in the uterus), although in up to 50% of cases, the tumor has a heterologous component (most commonly rhabdomyosarcoma or chondrosarcoma). There is no transition between the two components. Tumor stage is the most important prognostic factor in these tumors, although histologic features also affect

outcome. The finding of serous or clear-cell carcinoma is associated with a more aggressive course. Sarcomatous components adversely affect the overall prognosis of patients with stage I tumors (5-year survival is 30% among patients with heterologous elements as compared with 80% among patients with homologous elements); myometrial and lymphovascular invasion are also associated with a poor prognosis. Q: What are the treatment options for carcinosarcoma? A: Carcinosarcoma is thought to require multiple methods of treatment. Radiation therapy has been shown to reduce the rates of local recurrence in the pelvis but does not increase the survival benefit among patients with carcinosarcoma. Adjuvant chemotherapy has not been shown to have an effect on recurrence rates or progression-free or overall survival among patients with carcinosarcoma. Hormonal therapy is of no use, since estrogen and progesterone receptors do not control tumor growth, even though they are typically present in patients with carcinosarcoma.

QUOTE OF THE WEEK "In our study, more bleeding events of WHO grade 2, 3, or 4 occurred in the no-prophylaxis group than in the prophylaxis group, with a significant increase in the number of days with bleeding events of WHO grade 2, 3, or 4 and a decreased time to the first bleeding event of WHO grade 2, 3, or 4. Virtually all these bleeding episodes were WHO grade 2; only 7 of the 600 patients in the study had a bleeding event of WHO grade 3 or 4. More patients in the no-prophylaxis group had bleeding events of WHO grade 3 or 4, but this difference was not significant. The results of our study support the need for the continued use of prophylaxis . . . ." S.J. Stanworth and Others, Original Article, "A No-Prophylaxis