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Liver transplantation and de novo hepatitis B Infection
Liver transplantation and de novo hepatitis B Infection SiR-Chazouilleres and colleagues (Jan 15, p 142) present evidence of de novo acquisition in liver transplant recipients of hepatitis B virus (HBV) originating from the donor. Two such cases were confirmed by viral sequence homology, and in a further six the donor had antibodies to core (HBc) or surface (HBs) antigens. We present a further case, one which has
changed our donor assessment policy. The recipient was 57-year-old woman with primary biliary cirrhosis. Before transplantation she was HBsAg and anti-HBc negative. Liver transplantation was uneventful and involved the transfusion of blood products from 38 donors. The organ donor was HBsAg negative. The patient recovered well and was discharged with normal liver function tests. She presented 9 months post-transplant with a hepatic illness and was found to be positive for HBsAg, e antibody, and anti-HBc. All other virological testing was negative. Screening of stored serum revealed anti-HBc IgG 3 weeks after the transplant, this probably representing transient humoral graft-versus-host disease. Liver biopsy suggested acute viral hepatitis. There were no risk factors for hepatitis B after her transplant. Retrospective testing indicated that the infection had been acquired at the time of the operation, and confirmatory retrospective testing of all 38 blood donors was negative for HBsAg and anti-HBc. However, testing of serum from the liver donor indicated that she was positive for anti-HBc and anti-HBs. Polymerase chain reaction (PCR) testing for HBV core on serum and a liver sample taken 7 days post-transplant proved to be positive. The patient is now well with improving liver function tests. There have been other reports of de novo hepatitis B at the time of liver transplantation1.2 but few have been confirmed as donor in origin. In the UK, donors are screened for HBsAg but the inadequacy of this is demonstrated by our patient and those presented by Chazouilleres et al. Although de novo infection may be more benign clinically than recurrent hepatitis B, it is a potentially important cause of late morbidity. Our policy now is to test local donors for anti-HBc and to exclude, as donors of liver, any found positive. Testing donor blood for anti-HBc reduces the rate of transmission of HBV. Ideally all donors should be tested for HBV DNA but this is difficult to arrange. M A Aldersley, R E Eglin, J G Liver Unit, St James’s
O’Grady
University Hospital, Leeds LS9 7TF, UK
Wright T, Ferrell L, et al. Hepatitis C and B in liver transplantation. Transplant Proc 1993; 25: 2006-09. Douglas DD, Rakela J, Taswell HF, et al. Hepatitis B virus replication patterns after orthotopic liver transplantation: de novo versus recurrent infection. Transplant Proc 1993; 25: 1755-57. Takano S, Onata M, Ohto M, et al. Prospective assessment of donor blood screening for antibody to hepatitis C virus and high-titre antibody to HBcAg as a means of preventing posttransfusion hepatitis. Hepatology 1993; 18: 235—39.
1 Lake JR, 2
3
SIR-Chazouilleres and colleagues report that about 10% of patients in their programme express HBsAg post-transplant. Our experience in a paediatric liver transplantation programme is very different. Review of previous serology and retrospective analysis of stored sera was possible in 97 of 135 patients transplanted since 1984. None has been found to be HBsAg positive: 82 were negative 6 months or more post-transplant (range 6 months to 7 years), 9 were negative at 3-6 months, and 6 at 1-3 months. In the other 38 patients sera were unavailable but we have no reason to suppose that HBsAg positivity was more likely in those not assessed. The San Francisco group used PCR on DNA in stored samples to try and identify the source of infection. Were the
and storage stages handled in a routine diagnostic laboratory which regularly processes and stores samples from infected patients? The subsequent precautions would be negated by any prior contamination. We note that none of the PCR positive samples had other serological markers indicative of previous infection, and vice versa. Acquisition of HBV infection from health-care workers, notably surgeons, has been demonstrated1 and it would be useful to know what the local vaccination and screening policies were for those involved in invasive procedures. Cross-infection between patients is possible too and has been described in a setting where clinical trial volunteers had blood sampling via intravenous cannulae.2 Chazouilleres et al do not consider
separation
high-risk behaviour post-transplant (eg, intravenous drug abuse previously identified in 14 of 20 recipients) or sexual transmission. These two risk factors would not apply to a paediatric cohort and that difference might explain the difference between the two series.
Davison, R H George Microbiology Department and Liver Unit, Children’s Hospital, Birmingham B16 8ET, UK
D Mortiboy, S
1 2
Heptonstall J. Outbreak of hepatitis B virus infection associated with infected surgical staff. Commun Dis Rep 1991; 1: R81—85. Vickers J, Painter M, Heptonstall J, et al. Hepatitis B outbreak in a drug trials unit: investigation and recommendations. Commun Dis Rep 1994; 4: R1-4.
SIR-HBV infection can result in severe liver damage but little is known about what determines whether the infection becomes chronic or not. HLA class I restricted cytotoxic T-cells (CTL) may have a role.l-4 Chazouilleres and colleagues describe two cases of HBV infection in liver transplant recipients caused by "occult" HBV infection of the donor. After 38 and 56 months neither patient had cleared the virus. Here we describe an HBV-negative patient who received a liver from a chronic HBsAg carrier. However, despite the immunosuppression and a complete HLA class I mismatch, HBsAg was cleared from the infected organ and HBs antibodies developed within a year
(see table). A 57-year-old man, negative for all markers of hepatitis B or C virus infection was admitted in January, 1992, with end-stage alcoholic liver cirrhosis and hepatocellular carcinoma. 3 weeks
transplant was done. The blood groups of donor and recipient were identical but there was a complete HLA class I mismatch (donor A32, B52, B61, Cw2; recipient A2, A3, B7, B35, Cw4, Cw7). 3 days later the surgeons were told that the donor was a chronic HBsAg carrier. No special measures were taken, except for HBV specific immunohistology and serology. For immunosuppression a combination of cyclosporin, corticosteroids, and azathioprine was used. The patient recovered slowly and was discharged. During follow-up no complications attributable to HBV infection were observed and there were no histological signs of viral inflammation. After 150 days, an acute rejection was treated successfully with high-dose corticosteroids. Liver histology after 320 days showed a fatty liver, and the patient admitted alcohol abuse. The hepatocellular carcinoma has not recurred to date. later
a
Table: HBV markers In serum and liver before and after transplantation of HBsAg-positive liver
677
patient had no indication of HBV infection before transplantation but the first serum and liver samples, taken 3 days after transplantation, were HBsAg positive, confirming that the donor liver had been infected. The patient remained HBsAg positive in liver and serum for more than 200 days. There was no detectable HBeAg in the liver but at one time HBV-DNA was detected in serum. Thus the patient resembled a healthy HbsAg carrier. About 320 days after transplantation HBsAg was cleared from both liver and serum, and about 1 month later HBs antibodies developed. The patient is still anti-HBs positive. This case does not fit in with the view that CTLs are essential for the clearance of HBV for the patient’s effector cells should have been suppressed by the immunosuppressive therapy and unable to interact with the HLA-incompatible hepatocytes. Perhaps there are mechanisms
an enzyme immunosorbent were placed directly on In the other sera assay. laboratory, cells (Aedes albopictus mosquito cell line) and plaques C6/36 were sought after 14 days’ incubation. Dengue viruses were identified by IFA of cell monolayers by use of monoclonal antibodies.2 Culture fluids were tested by nested reverse transcriptase-polymerase chain reaction (RT-PCR) for dengue viruses.3 In addition, they were tested by plaque reduction neutralisation with serotype-specific polyclonal sera. Dengue viruses were recovered from 14 of 15 cases with acute flavivirus infection. Both methods of isolation were equivalent in recovery of viruses. Viral serotypes included dengue 2 (12 cases), dengue 3 (1 case), and mixed dengue 2/3 infection (1 case). The dual infection was evidenced by both IFA and RT-PCR; inoculation of T splendens yielded only one dengue
other than the classical antiviral CTL response which can terminate an HBV infection. In transgenic mice destruction of HBsAg-expressing liver cells can be mediated by cytokines, especially by interferon-gamma. If the CTLs responsible for the acute graft rejection in this patient also secreted interferongamma, this non-virus-specific immune response could have induced the clearance of HBV from the liver.
serotype (dengue 2). Our report documents the detection of dengue 3 virus in north-east Africa. Dengue 3 virus was originally isolated in Africa during an epidemic in Mozambique in 1985.4 Dengue serotypes 1 and 2 have previously been described in this region. Dengue viruses were responsible for 17% (14/81) of acute febrile illnesses in US forces in Somalia during this study. Dengue was the most common cause of fever in an endemic population from this region, in which 21 % showed evidence of infection. The appearance of dengue 3 in north-east Africa may reflect newly introduced virus or fortuitous recognition of circulating virus. The documented presence of three circulating serotypes of dengue virus may lead to the appearance of severe dengue (dengue haemorrhagic fever and dengue shock syndrome) in the region. We conclude that disease caused by dengue virus should be of increasing concern to medical personnel in north-east Africa. In addition, dengue infection should be considered in the evaluation of acute febrile illness in travellers returning from the region.
This
Supported by the Forschungsschwerpunkt Transplantation Heidelberg and grant from the Land Baden-Wurttemberg.
a
L Theilmann, J Arnold, U Töx, K Datsis, G Otto, W Hofmann, J Köck, H J Schlicht Department of Internal Medicine, University of Heidelberg, 69115 Heidelberg, Germany; Departments of Surgery, and Pathology, University of Heidelberg; and Department of Virology, University of Ulm 1 2
3
4
5
Waters J, Pignatelli M, Brown D, et al. The immune response to hepatitis B virus. Postgrad Med J 1987; 63 (suppl 2): 51-56. Bertoletti A, Ferrari C, Fiaccadori F, et al. HLA class I restricted human cytotoxic T cells recognize endogenously synthesized hepatitis B virus nucleocapsid antigen. Proc Natl Acad Sci 1991; 88: 10445-49. Missale G, Redeker A, Person J, et al. HLA-A31 and Aw68 restricted cytotoxic T cell responses to a single hepatitis B virus nucleocapsid epitope during acute viral hepatitis. J Exp Med 1993; 177: 751-62. Nayersina R, Fowler P, Guilhot S, et al. HLA A2 restricted cytotoxic T lymphocyte responses to multiple hepatitis B surface antigen epitopes during hepatitis B virus infection. J Immunol 1993; 150: 4659-71. Ando K, Moriyama T, Guidotti L, et al. Mechanisms of class I restricted immunopathology: a transgenic mouse model of fulminant hepatitis. J Exp Med 1993; 178: 1541-54.
dengue monoclonal antibodies in
Niranjan Kanesa-thasan, Lauren lacono-Connors, Alan Magill, Bonnie Smoak, David Vaughn, Doria Dubois, Jeanne Burrous, Charles Hoke Division of Communicable Diseases and Immunology and Division of Preventive Medicine, Walter Reed Army Institute of Research, Washington DC 20307-5100, USA; and Department of Virology, Armed Forces Research, Institute for Medical Sciences, Bangkok, Thailand
1 2
Dengue serotypes 2 and 3 in US forces In Somalia SIR-More than 25 000 US military personnel were stationed in Somalia during Operation Restore Hope in 1992-93. Personnel with fever were admitted to a central hospital facility. We did a prospective investigation of febrile illnesses between March and May, 1993, which revealed dengue viruses as an important identifiable cause of fever. 90 consecutively admitted patients with temperatures of at least 38’1°C were studied. Daily examinations and malaria smears were done; admission and discharge sera were collected and frozen in liquid nitrogen. Flavivirus infection was confirmed by positive dengue IgM and/or haemagglutination inhibition tests in 15 of 84 individuals tested. Positive serological tests were found only in cases of clinically suspected arboviral illness (abrupt onset of fever, severe headache, and myalgias without other illnesses). Arboviruses were sought in 81 available admission sera. Viral isolations were carried out at two separate laboratories. In one, acute sera were inoculated into Toxorhynchites splendens mosquitoes, and flaviviruses were identified with indirect immunofluorescence (IFA) of mosquito squash preparations by use of polyvalent flavivirus antibody.1 Viral isolates were identified with serotype-specific 678
3 4
5
Rosen L, Gubler DJ. The use of mosquitoes to detect and propagate dengue viruses. Am J Trop Med Hyg 1974; 23: 1153-60. Gubler DJ, Kuno G, Sather GE, Velez M, Oliver A. Mosquito cell cultures and specific monoclonal antibodies in surveillance for dengue viruses. Am J Trop Med Hyg 1984; 33: 158-65. Tanaka M. Rapid identification of flavivirus using the polymerase chain reaction. J Virol Methods 1993; 41: 311-22. Gubler DJ, Sather GE, Kuno G, Cabral JR. Dengue 3 virus transmission in Africa. Am J Trop Med Hyg 1986; 35: 1280-84. Hyams KC, Oldfield EC, Scott RM, et al. Evaluation of febrile patients in Port Sudan, Sudan: isolation of dengue virus. Am J Trop Med Hyg 1986; 35: 860-65.
Venodilation In
Raynaud’s disease
SiR-Bedarida and colleagues (Dec 11, p 1451) report no differences in venous responses to noradrenaline infusion in patients with Raynaud’s disease and controls. By contrast, we’ and others2 have shown significant group differences in vasoconstrictive responses to intra-arterial cx1-adrenergic and a2-adrenergic agonists when measuring total finger blood flow by venous occlusion plethysmography. Two points are noteworthy. First, total finger blood flow responses to intra-arterial drugs measure reactions of the whole digital vascular bed, whereas Bedarida’s method estimates only venous responses. In all likelihood, the vasospastic attacks of Raynaud’s disease are initiated in resistance, rather than
changed our donor assessment policy. The recipient was 57-year-old woman with primary biliary cirrhosis. Before transplantation she was HBsAg and anti-HBc negative. Liver transplantation was uneventful and involved the transfusion of blood products from 38 donors. The organ donor was HBsAg negative. The patient recovered well and was discharged with normal liver function tests. She presented 9 months post-transplant with a hepatic illness and was found to be positive for HBsAg, e antibody, and anti-HBc. All other virological testing was negative. Screening of stored serum revealed anti-HBc IgG 3 weeks after the transplant, this probably representing transient humoral graft-versus-host disease. Liver biopsy suggested acute viral hepatitis. There were no risk factors for hepatitis B after her transplant. Retrospective testing indicated that the infection had been acquired at the time of the operation, and confirmatory retrospective testing of all 38 blood donors was negative for HBsAg and anti-HBc. However, testing of serum from the liver donor indicated that she was positive for anti-HBc and anti-HBs. Polymerase chain reaction (PCR) testing for HBV core on serum and a liver sample taken 7 days post-transplant proved to be positive. The patient is now well with improving liver function tests. There have been other reports of de novo hepatitis B at the time of liver transplantation1.2 but few have been confirmed as donor in origin. In the UK, donors are screened for HBsAg but the inadequacy of this is demonstrated by our patient and those presented by Chazouilleres et al. Although de novo infection may be more benign clinically than recurrent hepatitis B, it is a potentially important cause of late morbidity. Our policy now is to test local donors for anti-HBc and to exclude, as donors of liver, any found positive. Testing donor blood for anti-HBc reduces the rate of transmission of HBV. Ideally all donors should be tested for HBV DNA but this is difficult to arrange. M A Aldersley, R E Eglin, J G Liver Unit, St James’s
O’Grady
University Hospital, Leeds LS9 7TF, UK
Wright T, Ferrell L, et al. Hepatitis C and B in liver transplantation. Transplant Proc 1993; 25: 2006-09. Douglas DD, Rakela J, Taswell HF, et al. Hepatitis B virus replication patterns after orthotopic liver transplantation: de novo versus recurrent infection. Transplant Proc 1993; 25: 1755-57. Takano S, Onata M, Ohto M, et al. Prospective assessment of donor blood screening for antibody to hepatitis C virus and high-titre antibody to HBcAg as a means of preventing posttransfusion hepatitis. Hepatology 1993; 18: 235—39.
1 Lake JR, 2
3
SIR-Chazouilleres and colleagues report that about 10% of patients in their programme express HBsAg post-transplant. Our experience in a paediatric liver transplantation programme is very different. Review of previous serology and retrospective analysis of stored sera was possible in 97 of 135 patients transplanted since 1984. None has been found to be HBsAg positive: 82 were negative 6 months or more post-transplant (range 6 months to 7 years), 9 were negative at 3-6 months, and 6 at 1-3 months. In the other 38 patients sera were unavailable but we have no reason to suppose that HBsAg positivity was more likely in those not assessed. The San Francisco group used PCR on DNA in stored samples to try and identify the source of infection. Were the
and storage stages handled in a routine diagnostic laboratory which regularly processes and stores samples from infected patients? The subsequent precautions would be negated by any prior contamination. We note that none of the PCR positive samples had other serological markers indicative of previous infection, and vice versa. Acquisition of HBV infection from health-care workers, notably surgeons, has been demonstrated1 and it would be useful to know what the local vaccination and screening policies were for those involved in invasive procedures. Cross-infection between patients is possible too and has been described in a setting where clinical trial volunteers had blood sampling via intravenous cannulae.2 Chazouilleres et al do not consider
separation
high-risk behaviour post-transplant (eg, intravenous drug abuse previously identified in 14 of 20 recipients) or sexual transmission. These two risk factors would not apply to a paediatric cohort and that difference might explain the difference between the two series.
Davison, R H George Microbiology Department and Liver Unit, Children’s Hospital, Birmingham B16 8ET, UK
D Mortiboy, S
1 2
Heptonstall J. Outbreak of hepatitis B virus infection associated with infected surgical staff. Commun Dis Rep 1991; 1: R81—85. Vickers J, Painter M, Heptonstall J, et al. Hepatitis B outbreak in a drug trials unit: investigation and recommendations. Commun Dis Rep 1994; 4: R1-4.
SIR-HBV infection can result in severe liver damage but little is known about what determines whether the infection becomes chronic or not. HLA class I restricted cytotoxic T-cells (CTL) may have a role.l-4 Chazouilleres and colleagues describe two cases of HBV infection in liver transplant recipients caused by "occult" HBV infection of the donor. After 38 and 56 months neither patient had cleared the virus. Here we describe an HBV-negative patient who received a liver from a chronic HBsAg carrier. However, despite the immunosuppression and a complete HLA class I mismatch, HBsAg was cleared from the infected organ and HBs antibodies developed within a year
(see table). A 57-year-old man, negative for all markers of hepatitis B or C virus infection was admitted in January, 1992, with end-stage alcoholic liver cirrhosis and hepatocellular carcinoma. 3 weeks
transplant was done. The blood groups of donor and recipient were identical but there was a complete HLA class I mismatch (donor A32, B52, B61, Cw2; recipient A2, A3, B7, B35, Cw4, Cw7). 3 days later the surgeons were told that the donor was a chronic HBsAg carrier. No special measures were taken, except for HBV specific immunohistology and serology. For immunosuppression a combination of cyclosporin, corticosteroids, and azathioprine was used. The patient recovered slowly and was discharged. During follow-up no complications attributable to HBV infection were observed and there were no histological signs of viral inflammation. After 150 days, an acute rejection was treated successfully with high-dose corticosteroids. Liver histology after 320 days showed a fatty liver, and the patient admitted alcohol abuse. The hepatocellular carcinoma has not recurred to date. later
a
Table: HBV markers In serum and liver before and after transplantation of HBsAg-positive liver
677
patient had no indication of HBV infection before transplantation but the first serum and liver samples, taken 3 days after transplantation, were HBsAg positive, confirming that the donor liver had been infected. The patient remained HBsAg positive in liver and serum for more than 200 days. There was no detectable HBeAg in the liver but at one time HBV-DNA was detected in serum. Thus the patient resembled a healthy HbsAg carrier. About 320 days after transplantation HBsAg was cleared from both liver and serum, and about 1 month later HBs antibodies developed. The patient is still anti-HBs positive. This case does not fit in with the view that CTLs are essential for the clearance of HBV for the patient’s effector cells should have been suppressed by the immunosuppressive therapy and unable to interact with the HLA-incompatible hepatocytes. Perhaps there are mechanisms
an enzyme immunosorbent were placed directly on In the other sera assay. laboratory, cells (Aedes albopictus mosquito cell line) and plaques C6/36 were sought after 14 days’ incubation. Dengue viruses were identified by IFA of cell monolayers by use of monoclonal antibodies.2 Culture fluids were tested by nested reverse transcriptase-polymerase chain reaction (RT-PCR) for dengue viruses.3 In addition, they were tested by plaque reduction neutralisation with serotype-specific polyclonal sera. Dengue viruses were recovered from 14 of 15 cases with acute flavivirus infection. Both methods of isolation were equivalent in recovery of viruses. Viral serotypes included dengue 2 (12 cases), dengue 3 (1 case), and mixed dengue 2/3 infection (1 case). The dual infection was evidenced by both IFA and RT-PCR; inoculation of T splendens yielded only one dengue
other than the classical antiviral CTL response which can terminate an HBV infection. In transgenic mice destruction of HBsAg-expressing liver cells can be mediated by cytokines, especially by interferon-gamma. If the CTLs responsible for the acute graft rejection in this patient also secreted interferongamma, this non-virus-specific immune response could have induced the clearance of HBV from the liver.
serotype (dengue 2). Our report documents the detection of dengue 3 virus in north-east Africa. Dengue 3 virus was originally isolated in Africa during an epidemic in Mozambique in 1985.4 Dengue serotypes 1 and 2 have previously been described in this region. Dengue viruses were responsible for 17% (14/81) of acute febrile illnesses in US forces in Somalia during this study. Dengue was the most common cause of fever in an endemic population from this region, in which 21 % showed evidence of infection. The appearance of dengue 3 in north-east Africa may reflect newly introduced virus or fortuitous recognition of circulating virus. The documented presence of three circulating serotypes of dengue virus may lead to the appearance of severe dengue (dengue haemorrhagic fever and dengue shock syndrome) in the region. We conclude that disease caused by dengue virus should be of increasing concern to medical personnel in north-east Africa. In addition, dengue infection should be considered in the evaluation of acute febrile illness in travellers returning from the region.
This
Supported by the Forschungsschwerpunkt Transplantation Heidelberg and grant from the Land Baden-Wurttemberg.
a
L Theilmann, J Arnold, U Töx, K Datsis, G Otto, W Hofmann, J Köck, H J Schlicht Department of Internal Medicine, University of Heidelberg, 69115 Heidelberg, Germany; Departments of Surgery, and Pathology, University of Heidelberg; and Department of Virology, University of Ulm 1 2
3
4
5
Waters J, Pignatelli M, Brown D, et al. The immune response to hepatitis B virus. Postgrad Med J 1987; 63 (suppl 2): 51-56. Bertoletti A, Ferrari C, Fiaccadori F, et al. HLA class I restricted human cytotoxic T cells recognize endogenously synthesized hepatitis B virus nucleocapsid antigen. Proc Natl Acad Sci 1991; 88: 10445-49. Missale G, Redeker A, Person J, et al. HLA-A31 and Aw68 restricted cytotoxic T cell responses to a single hepatitis B virus nucleocapsid epitope during acute viral hepatitis. J Exp Med 1993; 177: 751-62. Nayersina R, Fowler P, Guilhot S, et al. HLA A2 restricted cytotoxic T lymphocyte responses to multiple hepatitis B surface antigen epitopes during hepatitis B virus infection. J Immunol 1993; 150: 4659-71. Ando K, Moriyama T, Guidotti L, et al. Mechanisms of class I restricted immunopathology: a transgenic mouse model of fulminant hepatitis. J Exp Med 1993; 178: 1541-54.
dengue monoclonal antibodies in
Niranjan Kanesa-thasan, Lauren lacono-Connors, Alan Magill, Bonnie Smoak, David Vaughn, Doria Dubois, Jeanne Burrous, Charles Hoke Division of Communicable Diseases and Immunology and Division of Preventive Medicine, Walter Reed Army Institute of Research, Washington DC 20307-5100, USA; and Department of Virology, Armed Forces Research, Institute for Medical Sciences, Bangkok, Thailand
1 2
Dengue serotypes 2 and 3 in US forces In Somalia SIR-More than 25 000 US military personnel were stationed in Somalia during Operation Restore Hope in 1992-93. Personnel with fever were admitted to a central hospital facility. We did a prospective investigation of febrile illnesses between March and May, 1993, which revealed dengue viruses as an important identifiable cause of fever. 90 consecutively admitted patients with temperatures of at least 38’1°C were studied. Daily examinations and malaria smears were done; admission and discharge sera were collected and frozen in liquid nitrogen. Flavivirus infection was confirmed by positive dengue IgM and/or haemagglutination inhibition tests in 15 of 84 individuals tested. Positive serological tests were found only in cases of clinically suspected arboviral illness (abrupt onset of fever, severe headache, and myalgias without other illnesses). Arboviruses were sought in 81 available admission sera. Viral isolations were carried out at two separate laboratories. In one, acute sera were inoculated into Toxorhynchites splendens mosquitoes, and flaviviruses were identified with indirect immunofluorescence (IFA) of mosquito squash preparations by use of polyvalent flavivirus antibody.1 Viral isolates were identified with serotype-specific 678
3 4
5
Rosen L, Gubler DJ. The use of mosquitoes to detect and propagate dengue viruses. Am J Trop Med Hyg 1974; 23: 1153-60. Gubler DJ, Kuno G, Sather GE, Velez M, Oliver A. Mosquito cell cultures and specific monoclonal antibodies in surveillance for dengue viruses. Am J Trop Med Hyg 1984; 33: 158-65. Tanaka M. Rapid identification of flavivirus using the polymerase chain reaction. J Virol Methods 1993; 41: 311-22. Gubler DJ, Sather GE, Kuno G, Cabral JR. Dengue 3 virus transmission in Africa. Am J Trop Med Hyg 1986; 35: 1280-84. Hyams KC, Oldfield EC, Scott RM, et al. Evaluation of febrile patients in Port Sudan, Sudan: isolation of dengue virus. Am J Trop Med Hyg 1986; 35: 860-65.
Venodilation In
Raynaud’s disease
SiR-Bedarida and colleagues (Dec 11, p 1451) report no differences in venous responses to noradrenaline infusion in patients with Raynaud’s disease and controls. By contrast, we’ and others2 have shown significant group differences in vasoconstrictive responses to intra-arterial cx1-adrenergic and a2-adrenergic agonists when measuring total finger blood flow by venous occlusion plethysmography. Two points are noteworthy. First, total finger blood flow responses to intra-arterial drugs measure reactions of the whole digital vascular bed, whereas Bedarida’s method estimates only venous responses. In all likelihood, the vasospastic attacks of Raynaud’s disease are initiated in resistance, rather than