Association of type 2 diabetes mellitus and seroprevalence for cytomegalovirus.

Background: Infection is known to play a role in type 1 diabetes, but there is a paucity of information on its role in diabetes mellitus type 2. We examined the seroprevalence of selected viruses in a group of predominantly Hispanic patients with type 2 diabetes and control subjects without diabetes.

Methods: One hundred thirteen patients, 83 with type 2 diabetes and 30 control subjects, all undergoing hemodialysis at the same facility in San Antonio, Texas, were tested for antibodies against coxsackie B viruses, cytomegalovirus (CMV), and parvovirus. Sixty-six of these patients and 25 control subjects were tested bimonthly for 6 months.

Results: We observed a greater seroprevalence of anti-CMV IgG antibodies among patients with diabetes (97.6%), compared with control subjects (86.7%), and the difference was statistically significant [OR = 6.2, 95% CI: 1.1 to 36.0, P < 0.05]. Three draws on a subset of 91 patients produced still greater odds [OR = 12.4, 95% CI: 1.3 to 117, P < 0.05]. There were significantly more (P [less than or equal to] 0.001) vascular complications among patients with diabetes. There was a colinearity of trends between diabetes, seropositivity to CMV, and age.

Conclusions: Our findings indicated an up to 12 times greater odds of having type 2 diabetes for persons previously exposed to CMV. Since accelerated atherosclerosis is also associated with diabetes and CMV, past CMV infection may be a common factor that links atherosclerosis and diabetes. No other viruses tested in this study, either coxsackie B viruses or parvovirus, showed a significant association with type 2 diabetes.

Key Words: age, atherosclerosis, coxsackie B viruses, cytomegalovirus, hemodialysis, Mexican Americans, parvovirus, type 2 diabetes

**********

Type 2 diabetes is among the most prevalent chronic diseases worldwide. In 1997, the number of people with diabetes was estimated to be 135 million and, in 30 years, this is expected to increase to 300 million. (1) Numerous attempts have been made to identify causes of diabetes and its vascular complications. Among candidates are genetic factors, lipoprotein (a); advanced glycation end products; hyperglycemia; islet amyloid polypeptide; hyperhomocystinemia; clotting abnormalities; estrogen deficiency; and smoking. (2-10) However, it is estimated that the contribution of the most commonly measured risk factors cannot account for more than 5 to 25% of the excessive risk of vascular diseases and diabetes. (10,11) Thus, there are still other, unaccounted major risk factors that we must continue to explore. Among such unaccounted factors are inflammation and infection. (12,13) Past exposure to cytomegalovirus may be a common factor that links atherosclerosis and diabetes.

Research has been done using animal models to study the effect of viruses on pancreatic cells, (14) and several human studies addressed the role of viruses, cytomegalovirus (CMV), and coxsackie B viruses (CBV) in type 1 diabetes. (15-33) These studies involved either pancreatic analysis of a deceased diabetes patient or, more commonly, included the identification of elevated titers of CMV and CBV-neutralizing antibodies in type 1 diabetes patients, including newly diagnosed cases. Mertens et al (27) studied 166 patients with type 1 diabetes and found that 80% had neutralizing antibodies to CBV type B. He concluded that CBV infection occurred before the onset of diabetes. It has been estimated that approximately 50% of children with type 1 diabetes have antibodies to CBV. Barrett-Connor (34) and Toniolo et al (35) reported that one third of all type 1 diabetes cases are preceded by coxsackievirus type B infections.

In all, CBV and CMV have been linked with type 1 diabetes in studies that span more than three decades. However, little published work has been reported on the role viruses may play in type 2 diabetes. (36) To our knowledge, the present study is one of the first to address a possibility that infection may play a role in type 2 diabetes and its complications.

Materials and Methods

Eighty-three adults with type 2 diabetes and 30 control subjects without diabetes were enrolled in our study in January 1996, after obtaining informed consent. All subjects were undergoing hemodialysis three times per week in a chronic dialysis facility in San Antonio, Texas, and all used polysulfone high-flux F-80 artificial kidneys. Standard manual reuse processing was used for all dialyzers.

A case was defined as a dialysis patient with type 2 diabetes if he or she was (a) not insulin dependent; (b) had a known clinical history consistent with type 2 diabetes; and (c) had a diagnosis of type 2 diabetes by their nephrologist. Patients who had a history of diabetic ketoacidosis, were insulin dependent, or had an onset of their diabetes under age 20 were excluded. Control subjects were chronic dialysis patients who had unambiguous diagnoses of end-stage renal disease unrelated to diabetes (including chronic interstitial nephritis, primary hypertension, glomerulonephritis, or polycystic disease) and had no evidence of diabetic retinopathy.

Blood was obtained at the onset of the dialysis treatment and redrawn at 2-month intervals during the 6-month study period. Due to death, hospitalization, or transfer, only 91 (80%) of the total 113 participants, 66 cases and 25 comparison subjects, had samples drawn on all three occasions. Results were considered positive if at least one of three draws showed seropositivity to the specific virus. Fifteen milliliters of blood was drawn from each participant and analyzed for anti-CMV, CBV, and parvovirus (PVB) 19 antibodies (immunoglobulin [Ig]G and IgM). Both serum neutralization and indirect fluorescent immunoassay (IFA) with "Bion" reagents were used to test for CBV. (37) IFA was also used to test for CMV IgG and for CMV IgM, (38) using the Gull Laboratories, Inc., (Salt Lake City, UT) test kit. PVB19 was measured by IFA, (39) using an INCSTAR Corp. (Stillwater, MN) kit. All analyses were performed at the Virus Reference Laboratory in San Antonio, Texas. A sample was considered positive for anti-PVB19 IgG or IgM if a positive fluorescent result was obtained at a titer of 1:64 or greater (for IgG) and 1:16 or greater (for IgM). A sample was considered positive for serum neutralizing antibodies if a positive neutralization index was equal to or greater than 1:8 dilutions. A sample was considered positive for anti-CBV antibodies if the IFA test had a positive titer at 1:64 dilution (for IgG) and 1:8 for (IgM). A sample was considered positive for anti-CMV antibodies if the IFA test had a positive titer at 1:16 dilution (for IgG) and 1:10 (for IgM).

Patient information obtained from dialysis clinic records included age, sex, education, and socioeconomic status measured by Medicaid eligibility. Medical history information included number of months on dialysis, diagnoses, previous transfusions and renal transplants, as well as the type and extent of vascular complications (coronary and cerebrovascular disease, neuropathy, gastroparesis, retinopathy, hypertension, and amputations).

[chi square] and logistic regression analyses were performed, using SPSS software. (40) McNemar's matched analysis and power calculations were performed, using Epi-6 statistical software. (41) With 113 patients in total and a 50 to 60% prevalence of viruses of interest in the adult population, (42) our study was expected to detect, with a power of 80%, statistical odds of approximately 3 at a 95% confidence level. In the United States, prevalence of type 2 diabetes varies from 2 to 4% in whites, 4 to 6% in blacks, 10 to 15% in Hispanic Americans, and is as high as 35% in Native Americans.

Results

The patient population enrolled in this study consisted of 58 females and 55 males. One hundred subjects, or a predominant majority of participants, (84%) were Hispanic, with the remainder of the study group including five non-Hispanic white patients (4%), eight non-Hispanic black patients (7%), and one Asian patient (1%). Patients with diabetes, with a mean age of 62 years, were significantly older than referents, with a mean age of 49 years (P < 0.002). The study population was predominantly indigent (58%), using Medicaid eligibility as the criterion.

Serology results for the initial blood draws that were performed on 113 patients are presented in Table 1. After the first draw, 97.6% of patients with diabetes showed anti-CMV IgG antibodies, compared with 86.7% of control subjects, which produced a statistically significant odds ratio of 6.2 (P = 0.04). This difference remained significant in a subset of 91 patients which were available for all three draws (Table 2), with 98.5% CMV IgG-positivity among cases compared with 84.0% of control subjects. Having three draws increased the odds ratio for diabetes associated with seropositivity to CMV to OR = 12.4, 95% CI: 1.3 to 117. This odds ratio was statistically significant (P = 0.02) but had a larger confidence interval because of a smaller sample. Adjustment for patient age in a multiple logistic regression model removed the difference in the seropositivity to CMV (IgG).

Tables 3 and 4 show results of tests for acute infection. Eleven of 83 (13%) patients with diabetes had anti-CMV IgM antibodies at the beginning of this study (one draw), compared with 2 of 30 (7%) control subjects, producing an odds ratio equal to 2.4 (Table 3). In the subsequent 6 months, 14 of 66 (21%) patients with diabetes had at least one positive IgM test result out of three draws, compared with 3 of 25 (12%) control subjects (Table 4). Although the difference between cases and control subjects was not statistically significant, nearly twice as many patients with diabetes had acute infection with CMV that went unnoticed clinically.

There was no significant difference in seroprevalence of antibodies against other viruses, CBV and PVB19. However, 9 patients of 91 (10%) had an acute infection with PVB19 that also went undetected clinically.

Regardless of their status as cases or control subjects, patients with elevated IgG anti-CMV antibodies were on dialysis for an average of 38 months, and this was about one-half the time spent on dialysis by the CMV IgG-negative participants (77 months on average, P = 0.005). Therefore, the time spent in dialysis therapy had no relation to CMV-positive status. Also, there was no difference in the length of time spent on dialysis by participants with diabetes and control subjects. Similarly, there was no difference in the length of time spent on dialysis by CBV- and PVB19-positive and negative patients.

There were significantly more vascular complications among patients with diabetes, such as amputations (P = 0.001), retinopathy (P = 0.00001), neuropathy (P = 0.00006), gastroparesis (P = 0.00001), and cardiovascular disease (P = 0.00001), than in dialysis patients without diabetes.

Socioeconomic status, as measured by Medicaid eligibility and patient gender, were not significant predictors for diabetes or infection.

Discussion

In the United States, prevalence of type 2 diabetes varies from 2 to 4% in whites, 4 to 6% in blacks, to as much as 10 to 15% in Hispanic Americans, and even 35% in Native Americans. (43) According to McCulloch and Robertson, (9) whereas the incidence of new cases of type 2 diabetes seems fairly stable, the prevalence in the general population may be rising. The disease is generally asymptomatic for the first 4 to 7 years and might go unnoticed by the patient or doctor. (44) This is certainly true for Hispanic Americans (84% of patients in this study), who often are underinsured or have no health insurance and therefore have limited access to preventive health care. (45,46)

Evidence has linked viruses with the induction of insulin-dependent type 1 diabetes. Most studies suggested that CBV can cause diabetes by direct cytolysis of pancreatic beta cells, (14-16) whereas CMV was more likely to be associated with autoimmune diabetes, plus other factors, such as viral activation of the clotting cascade and alterations in the expressions of ELAM-1, ICAM-1, and VCAM-1 endothelial factors. (47) A strong correlation was reported by Foy et al (16) and Pak et al (17) between the genome of CMV and autoantibodies to islet cells found in the sera of insulin-dependent (type 1) diabetes.

That viruses may play a role in vascular disease has been extensively investigated. (12,13,47) Speir et al (48) examined 60 human coronary lesions and reported that there may be a significant relation between high amounts of the tumor suppressor protein p53 and human CMV. They proposed that angioplasty, performed for obstructed coronary arteries, may reactivate latent human CMV, with the release of a viral protein IE84 that blocks the inhibition of smooth muscle cell proliferation by p53. This mechanism is thought to lead to the development of coronary atherosclerosis. Since coronary artery disease is the most common cause of death both in persons with diabetes and end-stage renal disease, this observation may be significant to our study population. Our study revealed a very high prevalence of chronic CMV infection in patients with type 2 diabetes, as well as acute infection in nearly twice as many diabetic patients, compared with the control group.

Melnick et al (42,49) found CMV antigen and nucleic acid sequences in arterial smooth muscle cells, suggesting that CMV infection of the arterial wall may be common in patients with severe atherosclerosis. The high level of CMV antibodies was associated with clinically manifested vascular disease, possibly indicating a continuously active or periodically reactivated infection. By extension, one could expect that the accelerated vascular disease seen in diabetic patients on chronic dialysis might be related in part to latent CMV infection, manifested by severe atherosclerosis.

A recent article by French investigators (50) argued that a screen for genomic DNA from 18 carotid atherosclerotic lesions did not detect evidence of recent infection with CMV and other viruses. However, 9 (50%) of 18 specimens showed IgG antibodies for CMV, indicating that patients had a past infection with this virus. Authors concluded that this evidence does not rule out the possibility that infectious agents play an aggravating role in the development of atherosclerosis by increasing inflammation. In another study, German investigators (13) reported that elevated IgG antibodies were significantly associated with atherosclerosis (P < 0.05), adjusted for age, sex, and cardiovascular risk factors.

The present study was initiated to examine seroprevalence of antibodies to three viral agents, CMV, CBV, and PVB19, in a hemodialysis clinic population of end-stage renal disease patients. CMV and CBV were of interest because of their previous association with type 1 diabetes. PVB19 was included because it has not previously been associated with diabetes of either type and might serve as a reference or comparison virus.

We observed a high prevalence, up to 87%, of chronic CMV infection in dialysis patients without diabetes, but this proportion increased to as high as 98% in patients with type 2 diabetes. This difference was statistically significant, with a high odds ratio of up to 12 between cases with diabetes and control subjects. No other viruses tested in this study, either CBV or PVB19, showed a significant association with diabetes. There was an increased prevalence of PVB19 antibodies in patients with diabetes (92.7%), as compared with referents (83.3%), and this produced an elevated odds ratio (OR = 2.6). However, this odds ratio was not statistically significant at the 5% level test.

The seroprevalence of CMV infection is known to increase with age. In the United States, anti-CMV IgG antibodies have been reported in 10 to 15% of the adolescent population, 40 to 50% of the population by the age 35, and this may exceed 60 to 70% in the population older than 65. (36) Since the prevalence of type 2 diabetes also increases with age, (51) the relation between CMV, age, and diabetes mellitus type 2 may be confounded. Indeed, adjustment for age removed significant odds for CMV in our study population.

Vesseren et al (36) reported that CMV may play a role in the development of clinical atherosclerosis in patients with diabetes mellitus, even though age was shown to be a potentially confounding factor. Exactly by what mechanism age and diabetes are related is not well understood. However, both the prevalence of CMV and type 2 diabetes increase with age and both are associated with accelerated atherosclerosis.

In our study, we found an association between seropositivity for CMV and type 2 diabetes mellitus, but this may not necessarily mean there is a cause and effect relationship. Diabetic and dialysis patients both have decreased immunity, including cell-mediated immunity, and therefore it would be expected that they would be more susceptible to viral infections compared with patients without chronic renal failure and diabetes. Studies in the United States, Canada, and other countries reported greater prevalence of bacterial and viral illnesses, such as tuberculosis, influenza, hepatitis C virus, and others, in persons with diabetes, as compared with nondiabetic members of the same cohorts. (52-58)

This susceptibility to assorted infections in a population with compromised immunity would, at least in part, explain the possibility of increased CMV seropositivity in persons with type 2 diabetes, such as seen in our study. This infection might be latent and not clinically manifest.

Lohr and Oldstone (59) reported that latent CMV may cause no cytopathic effects and elicit no inflammatory response, yet the presence of this virus within cells can produce alterations in normal cellular functions, which might have pathologic consequences for the host organism. These authors examined preserved human pancreatic tissue samples from patients with type 2 diabetic and control subjects without diabetes for evidence of viral RNA. In the 32 patients with diabetes, no evidence was found for mumps, rubella, or CBV viruses. However, 14 (44%) of 32 pancreatic islet tissue samples from patients with diabetes mellitus type 2 were positive for CMV. Only one control subject of 50 (2%) was positive for CMV.

More recently, Gredmark et al (60) reported that CMV impairs immune functions by blocking certain aspects of cytokine-induced differentiation of monocytes, and this is a mechanism by which the virus escapes immune recognition and phagocytosis. The authors suggested that this may have clinical implication for the generalized suppression of immunity often observed in CMV-infected patients. This inhibition of macrophage differentiation was not observed in the same study for HIV and measles viruses.

In our study, not only was a very high prevalence of chronic CMV infection found in patients with type 2 diabetes, but also nearly twice as many patients with diabetes had acute infection, compared with control subjects. Overall, 17 participants, 14 of them with diabetes, of 91 patients who were tested three times during the 6-month study period, had evidence of acute CMV. Yet, no symptoms or signs of this infection were noted by their physicians. This observation is consistent with the findings by others, in that asymptomatic (occult) infection from a latent virus could be an unappreciated factor in diabetes or its vascular and infectious complications.

We cannot completely rule out a contribution of the dialysis process to the high prevalence of anti-CMV antibodies, but the likelihood is low. Viral antibodies are too large to pass through F-80 dialyzer membranes. (61) It may be that there was horizontal transmission of infection from patient to patient or from staff to patient. (46) However, the time spent on chronic dialysis, measured by the number of months, was shorter (by one-half) for patients with CMV infection than for patients without CMV.

Since transfused blood products can transmit CMV, (62) the number of transfusions received by patients with diabetes and control subjects were compared. There was no significant difference between the groups, but we were unable to exclude transfusions during previous hospitalizations for many of our study subjects.

Acknowledgment

We appreciate the input received from Kristina Mena, PhD, Assistant Professor of Virology, Division of Environmental and Occupational Health, UTSPH, El Paso Satellite Campus.

References

1. Vaaler S. Time for more effective secondary intervention. IDF Bull 1997;42:10-13.

2. Haffner SM. Lipoprotein (a) and diabetes. Diabetes Care 1993;16:835-840.

3. Brownlee M. Glycation and diabetic complications. Diabetes 1994;43:834-841.

4. Clark CM, Lee DA: Prevention and treatment of the complications of diabetes mellitus. N Engl J Med 1995;332:1210-1217.

5. Westermark P, Johnson KH, O'Brien TD, Betsholtz C. Islet amyloid polypeptide-a novel controversy in diabetes research. Diabetologia 1992;35.

6. Dennis VW, Robinson K. Homocysteinemia and vascular disease in end-stage renal disease. Kidney Int 1996;50:S11-S17.

7. Robinson JC, Folsom AR, Nabulsi AA, et al. Can postmenopausal hormone replacement improve plasma lipids in women with diabetes? The arteriosclerosis risk in communities study investigators. Diabetes Care 1996;19:480-485.

8. Wilson PW, Hoeg JM, D'Agostino RB, et al. Cumulative effects of high cholesterol levels, high blood pressure, and cigarette smoking on carotid stenosis. N Engl J Med 1996;337:516-522.

9. McCulloch DK, Robertson RP. Pathogenesis of type 2 diabetes mellitus. UpToDate www.uptodate.com 2003.

10. Horikawa Y, Oda N, Cox NJ. Genetic variation in the gene encoding calpain-10 is associated with type 2 diabetes mellitus. Nat Genet 2000;26:163.

11. Bierman E. Atherogenesis in diabetes. Arterioscler Thromb 1992;12:647-657.

12. Vercellotti GM. Overview of infection and cardiovascular diseases. J Allergy Clin Immunol 2001;108:S117-S119.

13. Espinola-Klein C, Rupprecht HJ, Blankenburg S, et al. Impact of infectious burden on extent and long-term prognosis of atherosclerosis. Circulation 2002;105:15-21.

14. Szopa TM, Titchener PA, Portwood ND, et al. Diabetes mellitus due to viruses-some recent developments. Diabetologia 1993;36:687-695.

15. Frisk G, Friman G, Tuvemo T, et al. Coxsackie B virus IgM in children at onset of type 1 (insulin-dependent) diabetes mellitus: evidence for IgM induction by a recent or current infection. Diabetologia 1992;35:249-253.

16. Foy CA, Quirke P, Williams DJ, et al. A search for candidate viruses in type 1 diabetic pancreas using the polymerase chain reaction. Diabetic Med 1994;11:564-569.

17. Pak CY, McArthur RG, Eun HM, et al. Association of cytomegalovirus infection with autoimmune type diabetes. Lancet 1988;2:1-4.

18. Gamble DR, Taylor KW, Cumming H. Coxsackie viruses and diabetes mellitus. Br Med J 1973;4:260-262.

19. Huff JC, Hierholzer JC, Farris WA. An "outbreak" of juvenile diabetes mellitus: consideration of a viral etiology. Am J Epidemiol 1974;100:277-287.

20. Anderson OO, Christy M, Amung K, et al. Viruses and Diabetes, in: Diabetes, Proceedings of the Ninth Congress of the International Diabetes Federation, in: Ninth Congress of the International Diabetes Federation, New Delhi, India, 1977, pp 294-298.

21. Schmidt WAK, Brade L, Munterfering H, et al. Course of Coxsackie B antibodies during juvenile diabetes. Med Microbiol Immunol (Berl) 1978;164:291-298.

22. Yoon JW, Austin M, Onodera T, et al. Virus-induced diabetes mellitus isolation of a virus from the pancreas of a child with diabetic ketoacidosis. N Engl J Med 1979;300:1173.

23. Hazra DK, Singh R, Wahal PK, et al. Coxsackie antibodies in young Asian diabetics. Lancet 1980;1:877.

24. Palmer JP, Cooney MK, Ward RH, et al. Reduced coxsackie antibody titres in type 1 (insulin-dependent) diabetic patients presenting during an outbreak of coxsackie B3 and B4 infection. Diabetology 1982;22:426-429.

25. Sakurami T, Nabeya N, Nagaoka K, et al. Antibodies to coxsackie B viruses and HLA in Japanese with juvenile-onset type 1 (insulin dependent) diabetes mellitus. Diabetology 1982;22:375-377.

26. King ML, Shaikh A, Bidwell D, et al. Coxsackie-B-virus-specific IgM responses in children with insulin-dependent (juvenile-onset; type 1) diabetes mellitus. Lancet 1983;1:1397-1399.

27. Mertens T, Gruneklee D, Hegges HJ. Neutralizing antibodies against coxsackie B viruses in patients with recent onset of type 1 diabetes. Eur J Pediatr 1983:140-293.

28. Prince MA, Vialettes B, Zevaco-Mattei C, et al. Clinical characteristics and etiological markers in insulin-dependent diabetes associated with an organ-specific autoimmune disease. Acta Diabetol Lat 1983;20:221.

29. Bell EJ, Mcartney RA. A study of coxsackie B virus infections 1972-1983. J Hyg Camb 1984;93:197.

30. Frisk G. Fohlman J, Kobbah M, et al. High frequency of coxsackie-B-virus-specific IgM in children developing type 1 diabetes during a period of high diabetes morbidity. J Med Virol 1985;17:219-227.

31. Frisk G, Nilsson E, Tuverno T, et al. The possible role of coxsackie A and echoviruses in the pathogenesis of type 1 diabetes mellitus studied by IgM analysis. J Infect 1992;24:13.

32. McCulloch DK. Pathogenesis of type 1 diabetes mellitus. UpToDate www.uptodate.com, 2003.

33. Foulis AK, McGill M, Farquharson MA, et al. A search for evidence of viral infection in pancreases of newly diagnosed patients with IDDM. Diabetologia 1997;40:53.

34. Barrett-Connor E. Is insulin-dependent diabetes mellitus caused by coxsackievirus B infection? A review of the epidemiologic evidence. Rev Infect Dis 1985;7:207.

35. Toniolo A, Federico G, Basolo F, et al. Diabetes Mellitus: Coxsackieviruses: A General Update. New York, Plenum Press, 1988.

36. Visseren FLJ, Bouter KP, Pon MJ, et al. Patients with diabetes mellitus and atherosclerosis, a role for cytomegalovirus? Diabetes Res Clin Pract 1997;36:49-55.

37. Lennette EH, Schmidt NJ (eds): Diagnostic Procedures for Viral, Rickettsial, and Chlamydial Infections. American Public Health Association, Washington, DC, 1979.

38. Hekker AC, Brand-Saathof B, Vis J, et al. Indirect immunofluorescence test for detection of IgM antibodies to cytomegalovirus. J Infect Dis 1979;140:596-600.

39. Brown CS, van Bussel MJA, Wassenaar ALM, et al. An immunofluorescence assay for the detection of parvovirus B19 IgG and IgM antibodies based on recombinant viral antigen. J Virol Methods 1990;29:53-62.

40. Norusis MJ. SPSS Advanced Statistics User's Guide. SPSS Inc., Chicago, 1989.

41. Dean AG, Dean JA, Coulombier D, et al. Epi Info Version 6. Atlanta, GA, Centers for Disease Control and Prevention, 1994.

42. Melnick JL, Adam E, Debakey ME. Cytomegalovirus and atherosclerosis. Eur Heart J 1993;14:30-38.

43. Bennett PH, Bogardus C, Tuomilehto J, et al. Epidemiology and natural history of NIDDM: non-obese and obese, in Alberti RA, DeFronzo H, Zimmet K, et al (eds): International Textbook of Diabetes Mellitus. John Wiley & Sons Ltd, Hoboken, 1992, pp 147-170.

44. Harris MI, Klein R, Welborn TA, et al. Onset of NIDDM occurs at least 4-7 yr before clinical diagnosis. Diabetes Care 1992;15:815-819.

45. Roberts B, Cech I. The Texas-Mexico border in 1993: important issues for the Clinton administration. San Antonio Med 1993;46:10-12.

46. Roberts B, Cech I, Alvarado NA. Diabetes educational programme in Texas: STOP reaches out. Diabetes Voice 1999;64:3-7.

47. Versallotti GM. Viruses and atherosclerosis: do they play a pathogenic role? J Invest Med 1998;46:403-407.

48. Speir E, Modali R, Huang E, et al. Potential role of human cytomegalovirus and P53 interaction in coronary restenosis. Science 1994;265:391-394.

49. Melnick JL, Hu C, Burek J, et al. Cytomegalovirus DNA in arterial walls of patients with atherosclerosis. J Med Virol 1994;42:170-174.

50. Watt S, Aesch B, Lanotte P, et al. Viral and bacterial DNA in carotid atherosclerotic lesions. Eur J Clin Microbiol Infect Dis 2003;22:99-105.

51. Jackson RA. Mechanisms of age-related glucose intolerance. Diabetes Care 1990;13:9-19.

52. Gu K, Cowie CC, Harris MI. Mortality in adults with and without diabetes in a national cohort of the US population, 1971-1993. Diabetes Care 1998;21:1138-1145.

53. Bertoni AJ, Saydah S, Brancati FL. Diabetes and the risk of infection-related mortality in the US. Diabetes Care 2001, 24:1044-1049.

54. Saxena AK, Panhotra BR. The susceptibility of patients with type 2 diabetes to hepatitis C virus infection during long-term haemodialysis. Swiss Med Wkly 2003;133:611-618.

55. Kawakami K. Tuberculosis in compromised hosts. Kekkaku 2003;78:717-722.

56. Shah BR, Hux JE. Quantifying the risk of infectious diseases for people with diabetes. Diabetes Care 2003;26:510-513.

57. Murray A, Foley R, Chavers B, et al. Infectious Complications of Hemodialysis, Peritoneal Dialysis, Transplant, and Chronic Kidney Disease Patients. United States Renal Data System Coordinating Center, Minneapolis, 2003.

58. Gerberding JL. Diabetes: Disabling, Deadly, and on the Rise. Atlanta, GA, Centers for Diseases Control and Prevention, 2004.

59. Lohr JM, Oldstone MB. Detection of cytomegalovirus nucleic acid sequences in pancreas in type 2 diabetes. Lancet 1990;336:644-648.

60. Grenmark S, Tilburg T, Soderberg-Naucler C. Human cytomegalovirus inhibits cytokine-induced macrophage differentiation. J Virol 2004;78:10378-10389.

61. Mujais SJ, Ivanovicj P. Membranes for extra corporeal therapy, in Maher JF (ed): Replacement of Renal Functions by Dialysis. Kluwer Academic Publishers, Boston, 1989.

62. Grundy JE, Ehrnst A, Einsele H, et al. A three-center European external quality control of study of PCR for detection of cytomegalovirus. J Clin Microbiol 1996;34:1166-1170.

This became a credo of mine ... attempt the impossible in order to
improve your work.
--Bette Davis

Bertram W. Roberts, MD, DRPH, and Irina Cech, PHD

From the School of Public Health, University of Texas, Houston, TX.

This study was conducted with the approval and in compliance with the research protocol submitted to the Committee for the Protection of Human Subjects. University of Texas Health Science Center at Houston.

The authors have no commercial or proprietary interests in the laboratory used for analysis of sera specimens, but Dr. Roberts was a former owner of Southwest Dialysis Center.

Reprint requests to Dr. Irina Cech, University of Texas School of Public Health, 1200 Herman Pressler, RAS E1005, Houston, TX 77030. Email: [e-mail]

Accepted December 16, 2004.

RELATED ARTICLE: Key Points

* Up to 12 times greater odds for having type 2 diabetes were observed for persons with elevated immunoglobulin G titers for cytomegalovirus, indicating previous exposure to this virus.

* Past exposure to cytomegalovirus may be a common factor that links atherosclerosis and diabetes.

Table 1. Seroprevalence of antibodies in patients with type 2 diabetes
and control subjects: Past infection, one draw

                                                              Diabetic
                                                              patients
                                                               n = 83
Virus                            Method        Results   Frequency  %

Cytomegalovirus (IgG)    Indirect fluorescent  Positive     81      97.6
                           assay               Negative      2       2.4
Coxsackie B virus (IgG)  Enzyme immunoassay    Positive     41      49.4
                                               Negative     42      50.6
Coxsackie B virus        Serum neutralization  Positive     12      14.5
  (neutralizing                                Negative     71      85.5
  antibodies)
Parvovirus B19 (IgG)     Indirect fluorescent  Positive     77      92.8
                           assay               Negative      6       7.2

                                    Nondiabetic
                                   control subjects
                                        n = 30       Significance
Virus                    Results   Frequency  %       (95% CI)

Cytomegalovirus (IgG)    Positive     26      86.7    OR = 6.2
                         Negative      4      13.3    (1.1-36.0)
                                                      P = 0.04
Coxsackie B virus (IgG)  Positive     18      60      OR = 0.6
                         Negative     12      40      (0.3-1.5)
                                                      P > 0.05
Coxsackie B virus        Positive      3      10.0    OR = 1.5
  (neutralizing          Negative     27      90.0    (0.4-5.8)
  antibodies)                                         P > 0.05
Parvovirus B19 (IgG)     Positive     25      83.3    OR = 2.6
                         Negative      5      16.7    (0.7-9.4)
                                                      P > 0.05

Table 2. Seroprevalence of viral antibodies in patients with type 2
diabetes and control subjects: Past infection, three draws

                                                            Diabetic
                                                            patients
                                                             n = 66
Virus                            Method        Results   Frequency  %

Cytomegalovirus (IgG)    Indirect fluorescent  Positive     65      98.5
                           assay               Negative      1       1.5
Coxsackie B virus (IgG)  Enzyme immunoassay    Positive     32      48.5
                                               Negative     34      51.5
Coxsackie B virus        Serum neutralization  Positive      9      13.6
  (neutralizing                                Negative     57      86.4
  antibodies)
Parvovirus B19 (IgG)     Indirect fluorescent  Positive     60      90.0
                           assay               Negative      6       9.1

                                     Nondiabetic
                                   control subjects
                                        n = 25       Significance
Virus                    Results   Frequency  %        (95% CI)

Cytomegalovirus (IgG)    Positive     21      84.0    OR = 12.4
                         Negative      4      16.0    (1.3-117)
                                                      P = 0.02
Coxsackie B virus (IgG)  Positive     15      60      OR = 0.6
                         Negative     10      40      (0.3-1.6)
                                                      P > 0.05
Coxsackie B virus        Positive      3      12.0    OR = 1.2
  (neutralizing          Negative     22      88.0    (0.3-4.7)
  antibodies)                                         P > 0.05
Parvovirus B19 (IgG)     Positive     19      76.0    OR = 2.6
                         Negative      6      24.0    (0.7-9.1)
                                                      P > 0.05

Table 3. Seroprevalence of viral antibodies in patients with type 2
diabetes and control subjects: Acute infection, one draw

                                                              Diabetic
                                                              patients
                                                               n = 83
Virus                           Method         Results   Frequency   %

Cytomegalovirus (IgM)    Indirect fluorescent  Positive      11     13.3
                           assay               Negative      72     86.7
Coxsackie B virus (IgM)  Enzyme immunoassay    Positive       1      1.2
                                               Negative      82     98.8
Parvovirus B19 (IgM)     Indirect fluorescent  Positive       1      1.2
                           assay               Negative      82     98.8

                                    Nondiabetic
                                  control subjects
                                       n = 30       Significance
Virus                    Results  Frequency   %       (95% CI)

Cytomegalovirus (IgM)    Positive      2       6.7    OR = 2.4
                         Negative     28      93.3    (0.5-10.3)
                                                      P > 0.05
Coxsackie B virus (IgM)  Positive      0       0      OR = 0.7
                         Negative     30      30      (0.1-6.5)
                                                      P > 0.05
Parvovirus B19 (IgM)     Positive      0       0      OR = 0.7
                         Negative     30      30      (0.1-6.5)
                                                      P > 0.05

Table 4. Seroprevalence of viral antibodies in patients with type 2
diabetes and control subjects: Acute infection, three draws

                                                             Diabetic
                                                             patients
                                                              n = 66
Virus                         Method         Results   Frequency  %

Cytomegalovirus (IgM)  Indirect fluorescent  Positive     14       21.2
                         assay               Negative     52       78.8
Coxsackie B virus      Enzyme immunoassay    Positive      0        0
  (IgM)                                      Negative     66      100.0
Parvovirus B19 (IgM)   Indirect fluorescent  Positive      7       10.6
                         assay               Negative     59       89.4

                                   Nondiabetic
                                control subjects
                                     n = 25       Significance
Virus                  Results  Frequency   %       (95% CI)

Cytomegalovirus (IgM)  Positive     3       12.0    OR = 2.0
                       Negative    22       88.0    (0.5-7.6)
                                                    P > 0.05
Coxsackie B virus      Positive     1        4.0    OR = 0.2
  (IgM)                Negative    25       96.0    (0.0-1.7)
                                                    P > 0.05
Parvovirus B19 (IgM)   Positive     2        8.0    OR = 1.4
                       Negative    23       92.0    (0.3-7.1)
                                                    P > 0.05