Immunological evidence of an early seroconversion to oncogenic Merkel cell polyomavirus in healthy children and young adults

Oncogenic Merkel cell polyomavirus (MCPyV) provokes a widespread and asymptomatic infection in humans. Herein, sera from healthy children and young adults (HC, n = 344) aged 0–20 years old were evaluated for anti-MCPyV immunoglobulin G (IgG) and IgM antibodies employing a recently developed immunoassay. Serum MCPyV IgG data from healthy subjects (HS, n = 510) and elderlies (ES, n = 226), aged 21–65/66–100 years old, from our previous studies, were included. The anti-MCPyV IgG and IgM rates in HC sera were 40.7% and 29.7%, respectively. A lower prevalence of anti-MCPyV IgGs was found in HC aged 0–5 years old (13%) compared to 6–10 (52.3%), 11–15 (60.5%) and 16–20 years old (61.6%) cohorts. Age-stratified HCs exhibited similar anti-MCPyV IgM rates (27.9%–32.9%). Serological profiles indicated that anti-MCPyV IgGs and IgMs had low optical densities (ODs) during the first years of life, while IgM ODs appeared to decrease throughout young adulthood. A lower anti-MCPyV IgGs rate was found in HC (40.7%) than HS (61.8%) and ES (63.7%). Upon the 5-years range age-stratification, a lower anti-MCPyV IgGs rate was found in the younger HC cohort aged 0–5 years old compared to the remaining older HC/HS/ES cohorts (52.3%–72%). The younger HC cohort exhibited the lowest anti-MCPyV IgG ODs than the older cohorts. Low anti-MCPyV IgMs rates and ODs were found in the 21–25 (17.5%) and 26–30 (7.7%) years old cohorts. Our data indicate that, upon an early-in-life seroconversion, the seropositivity for oncogenic MCPyV peaks in late childhood/young adulthood and remains at high prevalence and relatively stable throughout life.     

Antinuclear antibodies in the oldest-old women and men

The aim of this study was to compare the prevalence of antinuclear antibodies (ANA) in very old individuals (>or=86 years of age) with healthy younger (18-68 years) blood donors (n=200) regarding gender, health status, ratio of circulating CD4/CD8 cells and cytomegalovirus (CMV) serology. Frozen plasma was used for ANA detection in two study groups, i.e. 'OCTO' (97 persons aged 86-92 years, 65% women) and 'NONA' (136 persons aged 86-95 years, 70% women). OCTO participants were recruited on the basis that they were healthy or moderately healthy according to a selection protocol. No exclusion criteria regarding health status were applied in the NONA sample. The prevalence of ANA was significantly higher in the oldest-old samples compared to blood donors. There was no association between health status and the presence of ANA in the oldest-old. The difference across age was most pronounced in men, with low levels at younger age, whereas the prevalence among the oldest-old men reached similar levels as in women. There were no associations between the presence of ANA and CD4/CD8 ratio or with CMV status in the oldest-old. Our findings confirm an increased prevalence of ANA in the oldest-old, and emphasize the importance of taking gender and age into consideration when evaluating ANA.     

Reaction pattern of human anti-Mycoplasma pneumoniae antibodies in enzyme-linked immunosorbent assays and immunoblotting.

In serological diagnosis of Mycoplasma pneumoniae disease the frequently used complement fixation test is based on a cross-reacting glycolipid. Recently enzyme-linked immunoassays have been developed to overcome this lack of specificity. To study the involvement of the various proteins and the influence of age on the level of antiprotein antibodies present, we investigated by enzyme-linked immunoassays (immunoglobulin M [IgM] and IgG) and immunoblotting the sera of healthy persons of different age groups as well as sera of patients (including paired sera) with M. pneumoniae infection. In sera of children with nonrespiratory diseases and in healthy blood donors the IgM antibodies rose during the first 2 years of life to a relatively constant background level (optical density at 405 nm of 0.15 to 0.21). In contrast IgG remained low up to the seventh year and then increased to moderate levels (optical density of 0.15). The blotting patterns showed few IgM bands in the age group of 20 to 30 years. IgG blots revealed, up to 7 years, only very few reactions with a 168-kilodalton protein, but in higher age groups a considerable number of reactions with proteins of 193, 168, 84, 69, and 56 kilodaltons were detected. In the sera of patients, positive IgM blots were most numerous in the third week, whereas the number of IgG blots increased up to the fourth and the fifth week. At this time all sera contained antibodies against the 168-kilodalton protein, which is identical with the adhesin of M. pneumoniae. In a patient with acute infection, who had a high preinfection IgG level, no IgM response developed. The data indicate a relatively high background of antibodies against M. pneumoniae proteins in older age groups, suggesting a requirement for paired sera. Furthermore, reinfections of adults may occur without a concomitant IgM response.     

Antibody prevalence and specificity to group C rotavirus in Swedish sera

Of 160 sera collected from different age groups throughout Sweden, 38% were found to be antibody positive for group C rotavirus. The highest antibody prevalence rate was found in individuals aged 11-30 years (45%). An immunoprecipitation assay revealed that the antibodies were directed against VP2, VP4, VP6, VP7, and NSP2, with VP6 being the most immunogenic protein. Neutralising antibodies against a cultivable porcine group C rotavirus (strain AmC-1/Cowden) were detected in 16/19 individuals at titres from 160 to 5,120. The results indicate that group C rotavirus infections are relatively common in older Swedish children and adults but appear to be less common in children younger than 5 years of age. It is concluded that porcine and human group C rotaviruses share epitopes critical for stimulation of neutralising antibodies.     

Seroprevalence for toxoplasmosis in individuals living in North West Tuscany: access to Toxo-test in central Italy

This survey aimed to estimate the prevalence of anti-Toxoplasma IgG and IgM antibodies in people living in north west Tuscany (central Italy) and to investigate the adherence to antenatal screening programs and access to the Toxo-test as well. Sera from a large sample of individuals suspected to have acute infection or from pregnant women (10,352 subjects) aged between 1 day and over 70 years were analysed for both IgG and IgM anti-Toxoplasma antibodies using an immunoenzymatic method or a chemo-luminescent immunoassay. Overall, the seroprevalence of IgG antibodies was 21.4% (95% CI 20.62–22.20). A positive trend according to age was found, with low positivity observed in younger age groups. Among women of reproductive age the prevalence of IgG antibodies was 19.4% (95% CI 18.64–20.26). The overall IgM seroprevalence was 1.07% (95% CI 0.87–1.27). A low IgM prevalence was also observed in women of reproductive age (0.8%; 95% CI 0.65–1.03). Our study seems to indicate that primary prevention is widespread among women. However, an epidemiological surveillance system for toxoplasmosis should be implemented, to assess the risk of congenital toxoplasmosis and to determine the true burden of disease in adults.     

Evaluation of antibodies reactive with Campylobacter jejuni in Egyptian diarrhea patients.

Serum and stool samples were collected from 128 individuals: 96 diarrhea patients and 32 apparently healthy controls. Stool specimens were cultured for enteric bacterial pathogens, while sera were screened by enzyme-linked immunosorbent assay for Campylobacter jejuni-reactive antibodies. Of 28 diarrhea patients who demonstrated C. jejuni-reactive antibodies (titers, > 100), 14 were culture positive for this organism. The 32 healthy controls showed significantly lower antibody titers (P < 0.05) with the exception of 10 subjects who were culture positive for C. jejuni and had reactive immunoglobulin M (IgM) (6 subjects) and IgG (7 subjects). IgA was not detected in those 10 individuals (asymptomatic). Avidity was expressed as the thiocyanate ion concentration required to inhibit 50% of the bound antibodies. The avidity was higher in symptomatic patients than asymptomatic healthy controls. IgG was less avid (0.92 M) compared to IgM (0.1 M) and IgA (1.1 M), with no correlation between antibody titer and avidity. However, the thiocyanate ion concentration required for the complete inhibition of IgG (5 M)-bound antibodies was higher than that of IgA (2 M) and IgM (3 M). This study also shows that C. jejuni antibodies were variably cross-reactive with Escherichia coli, Shigella flexneri, Shigella sonnei, and Neisseria meningitidis in addition to Campylobacter coli and Campylobacter rectus.     

High prevalence of anti-cardiolipin and other autoantibodies in a healthy elderly population.

Serum samples from 64 apparently healthy individuals (32 men and 32 women, mean age 81.0 years) were examined for the prevalence of several autoantibodies, including rheumatoid factor (RF), antinuclear antibodies (ANA), antibodies to extracted cellular antigens Ro (SSA), La (SSB), Sm, U1nRNP and Scl-70. IgG and IgM isotype-specific ELISA methods were applied for the detection of antibodies to ssDNA (anti-ssDNA), to dsDNA (anti-dsDNA) and to cardiolipin (anti-CL). The sera of this elderly population were found to contain a plethora of autoantibodies; RF was detected in 14.1%, ANA in 31.3% and anti-Ro (SSA) in 1.6% of the individuals. Precipitating antibodies to La (SSB), Sm, U1nRNP and Scl-70 were absent, while 15.6% of the sera displayed precipitating antibodies to a common undefined human spleen antigen. ELISA methods revealed anti-ssDNA in 17.2% of the individuals, anti-dsDNA in 14.1% and anti-CL in an extremely high incidence (51.6%). Notably, the above autoantibodies were exclusively of IgG isotype. Tests of 261 sera from healthy non-elderly individuals disclosed only anti-CL (IgG and IgM isotypes) in 2.3% of them. The levels of IgA and IgG immunoglobulins were increased in 23.4% and 29.7% of the elderly subjects, respectively. IgM was elevated in 3.1%, but it was also found decreased in 9.4%. This study documents the high incidence of autoantibodies in the healthy elderly, including for the first time, anti-CL antibodies. Furthermore, the relative impairment in IgM autoantibody production observed, possibly indicates the involution of the senescent immune system.     

Specific IgG and IgA antibodies to herpes simplex virus (HSV)-induced surface antigen in patients with HSV infections and in healthy adults

Herpes simplex virus (HSV)-specific IgG and IgA antibody response in patients with HSV infection and in healthy adults was studied by the immunoperoxidase antibody-membrane antigen (IPAMA) technique. In all HSV infections in which specific IgM antibodies were detected by enzyme-linked immunosorbent assay (ELISA), a significant rise in the titer of HSV IgG and IgA antibodies was found. In contrast, in patients with recurrent herpes labialis in which no specific HSV IgM antibodies were detected by ELISA, HSV IgG and IgA antibodies were not found to fluctuate significantly during the course of infection. A higher geometric mean titer (GMT) for HSV IgG and for IgA antibodies was found in seropositive individuals with a previous history of recurrent HSV than in seropositive individuals without a previous history of recurrent HSV infection. Nineteen of 26 HSV IgG seropositive healthy medical students without a previous history of recurrent HSV infection had HSV IgA antibodies to membrane antigen. The significance of this finding in understanding the mechanism of latency in healthy seropositive individuals without previous history of HSV recurrent infections is discussed.     

Comparison of hepatitis A and E virus infections among healthy children in Mongolia: evidence for infection with a subgenotype IA HAV in children

To compare the epidemiologic profiles of hepatitis A virus (HAV) and hepatitis E virus (HEV) infections in children in Mongolia, the prevalence of HAV and HEV infections was investigated serologically and molecularly among 717 apparently healthy individuals of 0-20 years of age (mean +/- standard deviation, 8.6 +/- 4.9 years) using serum samples obtained between October 2005 and January 2006. Total antibody against HAV (anti-HAV [total]) was detected in 494 (68.9%) of the 717 subjects, while IgG antibody against HEV (anti-HEV IgG) was detected in only five subjects (0.7%) (P < 0.0001). All five subjects who had anti-HEV IgG, were negative for anti-HEV IgM and HEV RNA. Anti-HAV was detectable in 24 (75.0%) of the 32 infants aged 7 days to 6 months, but not in any of the 8 infants aged 7 to <12 months. The prevalence of anti-HAV was 19.5% (17/87) in the age group of 1-3 years, and it increased to 50.0% (69/138) in the age group of 4-6 years, and further to 81.4% (105/129) in the age group of 7-9 years. Of note, 97.2% of the subjects in the age group of 16-20 years had anti-HAV. The presence of HAV RNA was tested in all 717 subjects, and three children of 1, 4, or 8 years of age were found to have detectable HAV RNA (subgenotype IA). No subject had a history of hepatitis or jaundice. In conclusion, HEV infection was uncommon, but HAV infection lacking overt clinical features was prevalent among children in Mongolia.     

Seroprevalence of hepatitis E virus (HEV) in humans living in high pig density areas of Germany

An increase in acute autochthonous hepatitis E virus (HEV) infections has been recorded in Germany. These are suspected to be zoonotically transmitted from wild boar, deer and domestic pig. The latter may represent a major reservoir for HEV. In this study, 537 sera from humans living in Westphalia and Lower Saxony, representing areas of high pig density in Germany, were tested for the presence of HEV-specific antibodies. Among them were 302 individuals with occupational, direct contact to pigs and 235 individuals without direct contact to pigs. Two commercial tests and one in-house assay were applied for the detection of HEV-specific immunoglobulin G (IgG) antibodies. Sera were also tested in an assay that detects all classes of HEV-specific antibodies. Depending on the test used, the seroprevalence ranged from 4.1 to 27.9 %. Exposition to pigs was found to be associated with a significantly higher seroprevalence in subjects with contact to pigs (13.2–32.8 %) compared with that in non-exposed humans (7.7–21.7 %). In particular, individuals younger than 40 years with occupational exposure exhibited a markedly higher HEV seroprevalence compared with non-exposed individuals of that age group. In general, HEV seroprevalence increased with age resulting in a similar prevalence level in the age group of ≥50 years for exposed and non-exposed individuals. Analysis of all sera by a commercial anti-HEV IgM ELISA revealed 35 positive and 25 borderline samples. However, only one positive serum could be confirmed by an IgM line assay. Selected samples from IgM and/or IgG as well as total HEV antibody-positive individuals were also tested for the presence of HEV RNA. In one of the 78 samples, the only IgM ELISA positive and IgM line assay confirmed sample, RNA of HEV genotype 3 was detected. This sequence has high similarity to HEV sequences obtained from wild boars and domestic pigs from Germany and The Netherlands. This study demonstrates that in addition to the consumption of raw or undercooked meat, direct contact to pigs has to be considered as an additional risk factor for HEV infection.     

An Immunoblot Assay for Detection of Immunoglobulin M Antibody to Human Herpesvirus 6

We identified the human herpesvirus 6 (HHV-6)-dominant immunoglobulin M (IgM)-reactive virion protein as being the same 101-kDa protein (101K) previously identified as the major IgG immunoreactive protein and a specific serologic marker of HHV-6 infection. An immunoblot assay (IB) to detect HHV-6-specific IgM antibodies against the 101K protein in human serum samples was developed. The assay was validated by using acute- and convalescent-phase serum collected from children under 2 years of age in which we previously detected IgG seroconversion to the HHV-6 101K protein. Of 32 serum pairs which previously demonstrated IgG seroconversion to the 101K protein, 29 had IgM reactivity to the same protein in the acute-phase sample and the remaining 3 had reactivity in the convalescent-phase sample. We also detected HHV-6 IgM activity in sera collected from individuals ≥4 years of age who were also IgM seropositive to measles or rubella. Results of cross-adsorption studies using measles virus-, rubella virus-, and HHV-6-infected cells as the adsorbing antigen indicated no cross-reactivity between measles or rubella IgM and HHV-6 IgM in human serum samples. The IgM IB detected HHV-6-specific IgM antibody to the 101K protein in 78% (63 of 81) of tested acute-phase serum collected from young children with an undifferentiated rash illness by using a single serum dilution.     

Antibody responses to a C-terminal fragment of the Plasmodium falciparum blood-stage antigen Pf332 in Senegalese individuals naturally primed to the parasite

Previous studies have shown that antibodies from humans exposed continuously to malaria recognize the Plasmodium falciparum asexual blood-stage antigen Pf332. Here we analysed the antibody responses to a C-terminal fragment of Pf332, designated C231, in individuals from Senegal, by measuring the serum levels of immunoglobulin M (IgM), IgG class and subclass and IgE antibodies. IgG antibody reactivity with crude P. falciparum antigen was detected in all the donors, while many of the children lacked or had low levels of such antibodies against C231. The antibody levels increased significantly with age for both crude P. falciparum antigen and C231, and in the older age groups most of the donors displayed antibodies to C231. This was also true for IgM, IgE and IgG subclass reactivity against C231. Moreover, the ratio of IgG1/IgG2 was considerably lower for C231 than for crude P. falciparum antigen, and in age groups 10-14 and 15-19 years the levels of IgG2 against C231 even exceeded that of IgG1. The IgG2/IgG3 ratios suggest that C231 gives similar levels of IgG2 and IgG3, except for children aged 4-9 years, where IgG3 was higher. Raw IgM, IgG class and subclass and IgE antibody levels to C231 tended to be higher in those who did not experience a malaria attack, but following linear multivariate analysis the trends were not significant.     

Antinuclear antibodies in the oldest-old women and men

The aim of this study was to compare the prevalence of antinuclear antibodies (ANA) in very old individuals (≥86 years of age) with healthy younger (18–68 years) blood donors (n = 200) regarding gender, health status, ratio of circulating CD4/CD8 cells and cytomegalovirus (CMV) serology. Frozen plasma was used for ANA detection in two study groups, i.e. ‘OCTO’ (97 persons aged 86–92 years, 65% women) and ‘NONA’ (136 persons aged 86–95 years, 70% women). OCTO participants were recruited on the basis that they were healthy or moderately healthy according to a selection protocol. No exclusion criteria regarding health status were applied in the NONA sample. The prevalence of ANA was significantly higher in the oldest-old samples compared to blood donors. There was no association between health status and the presence of ANA in the oldest-old. The difference across age was most pronounced in men, with low levels at younger age, whereas the prevalence among the oldest-old men reached similar levels as in women. There were no associations between the presence of ANA and CD4/CD8 ratio or with CMV status in the oldest-old. Our findings confirm an increased prevalence of ANA in the oldest-old, and emphasize the importance of taking gender and age into consideration when evaluating ANA.     

IgM, IgG and IgG subclass antibodies to herpes simplex virus in persons of different ages

IgM, IgG antibodies to herpes simplex virus and their subclases were investigated in 565 subjects of different age tested at virological laboratories of St. Petersburg in 1996-1997. The majority of these subjects had a history of herpes infection and 21.5% had IgM antibodies to herpes simplex virus (HSV), marker of acute herpetic infection. Besides IgM, IgG1 antibodies can be referred to early antibodies appearing during the acute stage of herpetic infection. The predominant subclass was HSV IgG3 antibodies. As for IgG4, they were completely absent in infants aged under 1 year, were detected in 6.2% children aged under 14 years, and were present in 12.2-12.5% adults.     

Increased risk of parvovirus B19 infection in young adult cancer patients receiving multiple courses of chemotherapy

An increased human parvovirus B19 infection rate has been observed in immunocompromised hosts. In this study, we sought to determine the prevalence of parvovirus B19 infection in adult cancer patients receiving multiple courses of systemic chemotherapy. From March 1999 through April 2000, 59 men and 68 women, with a median age of 49 (18 to 79) years, were enrolled in this study. They had received an average of 7.1 (4 to 32) courses of systemic chemotherapy. The median duration from the date of starting chemotherapy to the date of blood sampling was 11 (4 to 88) months. Serum B19 immunoglobulin G (IgG) and IgM levels were examined by an enzyme-linked immunosorbent assay, and B19 DNA was examined by a nested PCR. A group of 400 healthy blood donors served as the control group. The overall prevalences of anti-B19 IgG in adult cancer patients and healthy blood donors were 61.4 and 25.0%, respectively (P < 0.01). Anti-B19 IgM and B19 DNA were not detectable in these anti-B19 IgG-seropositive individuals. A further age-stratified comparison revealed that only patients younger than 40 years had a significantly higher anti-B19 IgG seropositivity rate than the controls (19 of 39 versus 53 of 310; P < 0.001). The increased prevalence of B19 infection in these 39 adult patients younger than 40 years might be clinically significant, since unexplained anemia, a common sequela of B19 infection, was detected in 3 of 20 seronegative patients (15.0%) and in 12 of 19 seropositive patients (63.2%) (P < 0.005). The results of this study suggest that adult patients younger than 40 years and receiving multiple courses of systemic chemotherapy may have a significantly increased risk of B19 infection. Prospective studies to define the time course and clinical consequence of B19 infection in this group of patients are needed.     

A nationwide survey of hepatitis E virus infection in the general population of Japan

To investigate nationwide the prevalence of hepatitis E virus (HEV) infection in the general population of Japan, serum samples were collected from 22,027 individuals (9,686 males and 12,341 females; age, mean ± standard deviation: 56.8 ± 16.7 years; range: 20–108 years) who lived in 30 prefectures located in Hokkaido, mainland Honshu, Shikoku, and Kyushu of Japan and underwent health check‐ups during 2002–2007, and were tested for the presence of IgG, IgM, and IgA classes of antibodies to HEV (anti‐HEV) by in‐house ELISA and HEV RNA by nested RT‐PCR. Overall, 1,167 individuals (5.3%) were positive for anti‐HEV IgG, including 753 males (7.8%) and 414 females (3.4%), the difference being statistically significant (P < 0.0001). The prevalence of anti‐HEV IgG generally increased with age and was significantly higher among individuals aged ≥50 years than among those aged <50 years (6.6% vs. 2.7%, P < 0.0001). Although 13 individuals with anti‐HEV IgG also had anti‐HEV IgM and/or anti‐HEV IgA, none of them had detectable HEV RNA. The presence of HEV RNA was further tested in 50 or 49‐sample minipools of sera from the remaining 22,014 individuals, and three individuals without anti‐HEV antibodies tested positive for HEV RNA. The HEV isolates obtained from the three viremic individuals segregated into genotype 3 and were closest to Japan‐indigenous HEV strains. When stratified by geographic region, the prevalence of anti‐HEV IgG as well as the prevalence of HEV RNA or anti‐HEV IgM and/or anti‐HEV IgA was significantly higher in northern Japan than in southern Japan (6.7% vs. 3.2%, P < 0.0001; 0.11% vs. 0.01%, P = 0.0056; respectively). J. Med. Virol. 82:271–281, 2010. © 2009 Wiley‐Liss, Inc.     

Frequency of anti-hsp60, -65 and -70 antibodies in sera of patients with juvenile idiopathic arthritis

Cross-reactivity between microbial and human heat shock proteins (hsps) led to the concept that hsp might be involved in the etiopathogenesis of autoimmune diseases. We investigated antibodies to recombinant human hsp60, recombinant Mycobacterium bovis hsp65 and to stress-inducible recombinant human hsp70 using enzyme-linked immunosorbent assay (ELISA) in sera of 209 juvenile idiopathic arthritis (JIA) patients and 50 healthy controls. Anti-hsp60 antibodies did not exceed the control level in any JIA patient. The numbers of JIA patients (16/209, 7.6%) who raised anti-hsp65 antibodies was equal to healthy controls (4/50, 8%). Elevated levels of antibodies against hsp70 were found in a cohort of patients with JIA (36.8%) when compared with age-matched healthy individuals (2%). These antibodies were predominantly of IgG isotype in systemic disease and IgM isotype in oligoarthritis. In polyarthritis both IgG and IgM antibodies frequently occurred. Significantly higher anti-hsp70 antibody levels were found in RF-positive JIA patients. The levels of anti-hsp70 antibodies correlated with the severity of disease evaluated on the basis of Steinbrocker's functional classification and rtg staging system. No association between anti-hsp70 antibody levels and ANA, HLA B27 and disease duration (less than 2 years x more than 2 years) was observed except IgM anti-hsp70 antibody where significantly higher levels were also detected in HLA B27-positive patients. The prevalence of anti-hsp70 antibodies is much higher in JIA patients when compared with healthy controls, suggesting their possible role in pathological mechanism of the disease.     

Seroprevalence of Toxoplasma gondii in northern Greece during the last 20 years

The seroprevalence of Toxoplasma gondii in the northern Greek population was determined in 1984, 1994 and 2004, and changes during this period were investigated. In total, 1014, 812 and 958 sera from individuals aged 1 day to 70 years were examined in 1984, 1994 and 2004, respectively, for IgG and IgM anti-Toxoplasma antibodies with the standard immunofluorescence assay (IFA) and microparticle enzyme immunoassay (MEIA). In individuals positive for IgM-specific antibodies, primary infection with Toxoplasma was diagnosed on the basis of the Toxoplasma serological profile (IFA, MEIA, conventional IgM and IgA ELISAs, immunosorbent agglutination assay and IgG avidity test). The prevalence of IgG-specific antibodies in the general population was 37%, 29.9% and 24.1% in 1984, 1994 and 2004, respectively, and was 35.6%, 25.6% and 20%, respectively, in women of reproductive age (15-39 years). The incidence of Toxoplasma infection, based on cases of primary infection and the annual seroconversion rate for the general population, was estimated to be 1.25% and 1.1% in 1984, 1.05% and 0.93% in 1994, and 0.85% and 0.8% in 2004. The significant decline in prevalence, and the shift towards an older age group, observed during this period could be explained by the improved socio-economic situation. The high (80%) proportion of women of reproductive age susceptible to Toxoplasma infection, with an estimated 90-200 neonates infected in utero annually, seems to present a potential risk to public health. Education of the public and prophylactic measures may become increasingly important.     

Enzyme-linked immunosorbent assay for immunological diagnosis of human tularemia.

The enzyme-linked immunosorbent assay (ELISA) was applied for immunological diagnosis of human tularemia, using lipopolysaccharide from Francisella tularensis as antigen. Sera collected from patients, healthy individuals, and vaccinated volunteers were investigated for antibodies against F. tularensis by ELISA and tube agglutination. In ELISA all sera were titrated with a polyspecific anti-immunoglobulin enzyme conjugate. A limited number of consecutive sera from individual patients were also investigated for immunoglobulin G (IgG) and IgM antibodies by means of immunoglobulin class-specific conjugates. On an average ELISA was more than 10-fold as sensitive as tube agglutination. Two weeks after onset of disease, sera from patients had significantly higher titers in ELISA than sera from healthy controls. High titers persisted after more than 2 years. Significant amounts of both IgG and IgM antibodies were present within 1 to 2 weeks after infection. The antibody activity increased during the first month, without any significant change of the relation between IgG and IgM titers. After 2.5 years the IgG/IgM titer ratio of sera from patients was significantly increased. Within 6 weeks after vaccination sera from about half of the vaccinees had significantly elevated titers in ELISA. Titers observed after vaccination were generally lower than those found after infection. An elevated ELISA titer can be of diagnostic importance by the end of the first week of illness. A significant increase of titer in consecutive serum samples indicates a diagnosis of tularemia. Determination of IgG and IgM antibodies may be of value in determing whether a positive titer of a single serum sample is of longstanding or recent origin.     

An immunoblot assay for detection of immunoglobulin M antibody to human herpesvirus 6

We identified the human herpesvirus 6 (HHV-6)-dominant immunoglobulin M (IgM)-reactive virion protein as being the same 101-kDa protein (101K) previously identified as the major IgG immunoreactive protein and a specific serologic marker of HHV-6 infection. An immunoblot assay (IB) to detect HHV-6-specific IgM antibodies against the 101K protein in human serum samples was developed. The assay was validated by using acute- and convalescent-phase serum collected from children under 2 years of age in which we previously detected IgG seroconversion to the HHV-6 101K protein. Of 32 serum pairs which previously demonstrated IgG seroconversion to the 101K protein, 29 had IgM reactivity to the same protein in the acute-phase sample and the remaining 3 had reactivity in the convalescent-phase sample. We also detected HHV-6 IgM activity in sera collected from individuals > or =4 years of age who were also IgM seropositive to measles or rubella. Results of cross-adsorption studies using measles virus-, rubella virus-, and HHV-6-infected cells as the adsorbing antigen indicated no cross-reactivity between measles or rubella IgM and HHV-6 IgM in human serum samples. The IgM IB detected HHV-6-specific IgM antibody to the 101K protein in 78% (63 of 81) of tested acute-phase serum collected from young children with an undifferentiated rash illness by using a single serum dilution.