Recognition of different Toxoplasma antigens by IgM and IgG antibodies in mothers and their congenitally infected newborns

The protein-blotting technique was used to determine the antigens of Toxoplasma gondii that were recognized by IgG and IgM antibodies in sera of congenitally infected newborns and their mothers. Patterns of IgG and IgM blots with sera from newborns revealed antigen-antibody reactions (bands) that were not present in the respective blots obtained with sera from their mothers. This was true for 50% of 24 congenitally infected newborns. In contrast, such a difference was noted in only one (5%) of 21 newborns who were not congenitally infected but whose mothers had serological evidence of acute infection with T. gondii acquired during gestation. Our results suggest that the protein-blotting method or an adaptation may be valuable for study of the immune response of the mother, fetus, and newborn to various antigens of infectious organisms and for diagnosis of congenital infections in the newborn.     

Human fetuses are able to mount an adultlike CD8 T-cell response

Fetal/neonatal immune responses generally are considered to be immature and weaker than that of adults. We have studied the cord-blood T cells of newborns congenitally infected with Trypanosoma cruzi, the protozoan agent of Chagas disease. Our data demonstrate a predominant activation of CD8 T cells expressing activation markers and armed to mediate effector functions. The analysis of the T-cell receptor beta chain variable repertoire shows the oligoclonal expansion of these T lymphocytes, indicating that activation was driven by parasite antigens. Indeed, we have detected parasite-specific CD8 T cells secreting interferon-gamma after coincubation with live T cruzi. This response is enhanced in the presence of recombinant interleukin-15, which limits the T-cell spontaneous apoptosis. These findings point out that the fetal immune system is more competent than previously appreciated, since fetuses exposed to live pathogens are able to develop an adultlike immune CD8 T-cell response.     

Are maternal re‐infections with Trypanosoma cruzi associated with higher morbidity and mortality of congenital Chagas disease?

BACKGROUND: Comparing two surveys performed in Bolivia in 1992-1994 and 1999-2001, we reported a significant decrease in the proportions of severe and mortal forms of congenital Chagas disease. This might be due to a reduction of vectorial density (VD) in maternal residence area, raising the question of a possible causal relationship between such VD, maternal parasitaemia and prognosis of congenital infection with Trypanosoma cruzi. METHOD: Comparisons of haematological and parasitological data obtained from Bolivian mothers infected with T. cruzi, and of clinical and biological data obtained from their infected and uninfected newborns, stratified according to VD in the area of maternal residence. RESULTS: i) Blood hematocrit rates or hemoglobin amounts were within the normal ranges and similar in all the maternal groups, whatever the VD in their areas of residence; ii) mothers living in high VD areas displayed a higher frequency of hemocultures positive for T. cruzi; iii) newborns congenitally infected with T. cruzi, but not uninfected babies born from infected mothers, displayed higher frequencies of very low Apgar scores, low birth weights, prematurity, respiratory distress syndrome or anasarca, as well as higher mortality rates when their mothers lived in areas of high VD. CONCLUSION: Frequent bites of blood sucking Reduvidae during pregnancy do not induce maternal anaemia, but, likely through multiple maternal re-infections with T. cruzi, increase maternal parasitemia and worsen congenital Chagas disease. Maternal dwelling in areas of high VD is associated with a serious increased risk of severe and mortal congenital Chagas disease.     

Laser-capture microdissection of plasma cells from subacute sclerosing panencephalitis brain reveals intrathecal disease-relevant antibodies

Increased IgG and oligoclonal bands are found in cerebrospinal fluid of humans with chronic infectious CNS disease. Studies have shown that these oligoclonal bands are antibodies directed against the agent that causes disease. Laser-capture microdissection was used to isolate individual CD38+ plasma cells from the brain of a patient with subacute sclerosing panencephalitis, and single-cell RT-PCR was used to analyze individual IgG heavy and light chains expressed by each cell. Based on overrepresented IgG sequences, we constructed functional recombinant antibodies (recombinant IgGs) and determined their specificities. Five of eight recombinant IgGs recognized measles virus, the cause of subacute sclerosing panencephalitis. These results demonstrate that overrepresented IgG sequences in postmortem brains can be used to produce functional recombinant antibodies that recognize their target antigens. This strategy can be used to identify disease-relevant antigens in CNS inflammatory diseases of unknown etiology.     

Antibody drop in newborns congenitally infected by Trypanosoma cruzi treated with benznidazole

Objective  To compare the drop of Chagas antibody titres between non-infected and congenitally infected newborns treated by two doses of benznidazole, aiming at evaluating the recovery time and giving recommendations regarding serological criteria of recovery.
Methods  During a clinical trial, the drop of Trypanosoma cruzi antibody titres measured by ELISA tests was followed during the first year of life in congenitally infected newborns treated with different doses of benznidazole and compared to T. cruzi antibody titres in non-parasitaemic newborns. Confirmation of recovery was given by two negative serological tests: Chagas Stat-Pak^® (CSP) (immunochromatography) and Chagatest^® v3.0 (ELISA).
Results  In non-parasitaemic infants of infected mothers, antibodies of maternal origin disappeared in <8 months while in infected infants, T. cruzi antibodies decreased more slowly and disappeared in 9–16 months allowing to confirm the recovery. All CSP tests were negative before the ninth month while about 10% of ELISA tests remained positive at the 12th month.
Conclusions  Recovery may be confirmed in most cases at 10 months. The CSP test was compared to Chagatest^® v3.0 ELISA and appeared to give a reliable response. The decrease rate of antibodies does not depend on treatment modes.     

Recognition of Treponema pallidum antigens by IgM and IgG antibodies in congenitally infected newborns and their mothers

Immunoblotting techniques were used to examine the proteins of Treponema pallidum recognized by IgM and IgG antibodies in sera from infants with congenital syphilis and their mothers. Infected infants' serum IgM reactivity to treponemal antigens differed from that of control infants born to normal, serofast, and biologic false-positive mothers. Each of the infected infants' sera exhibited IgM reactions to the 47- and 37-kilodalton (kDa) proteins of T. pallidum. Although rheumatoid factor was detected in the sera of half of the infected infants, removing this factor did not alter the pattern of IgM blots. IgG reactions in infants were almost exclusively of the IgG1 and IgG3 subclasses and mirrored those of the mother, except for IgG1 and IgG3 reactions to the 83-kDa treponemal protein, which were unique to infants' sera. Our results suggest that the findings of IgM antibody directed against the 47- or 37-kDa antigens of T. pallidum may help to diagnose congenital syphilis at birth.     

Immunodetection of antibodies in sera from symptomatic and asymptomatic Chilean Chagas' disease patients with Trypanosoma cruzi recombinant antigens.

A panel of eight Trypanosoma cruzi antigens produced by recombinant DNA techniques was used to compare the reactivity of IgG specificities in the sera from 45 chronic Chagas' disease patients with different clinical symptoms (cardiac disease, gastrointestinal lesions, and combined syndrome) with those present in the sera from 55 asymptomatic patients in Chile. All of the serum samples were first characterized for antibody to T. cruzi epimastigotes by immunofluorescence assay. All of the Chagas' disease sera were reactive, but none of five healthy controls whose sera were also tested had antibodies against the fixed parasites. A dot-blot assay was then performed to evaluate the serum reactivity against recombinant DNA clones 1, 2, 13, 26, 30, 36, 54, and SAPA (shed acute phase antigen). These recombinant antigens were recognized by a large proportion of the sera collected from the Chilean patients. Ninety-five percent of the serum samples reacted with one or more of the recombinant clones. Analysis of the reactivity with individual fusion proteins showed that 88% of these sera reacted with clones 1 and 2, and 78% reacted with clone 13. Differences in reactivity to clones number 13, 30, and SAPA were observed when symptomatic and asymptomatic patients were compared. These differences in reactivity were statistically significant (P less than 0.01) according to Fisher's exact test.     

Molecular analysis of the fetal IgM response to Treponema pallidum antigens: implications for improved serodiagnosis of congenital syphilis

Western blot analysis of the fetal IgM response to Treponema pallidum antigens was examined among 39 pairs of maternal/infant sera; this included 12 mothers and infants with active syphilis (group I), 9 mothers with active syphilis and their infants with uncertain infection (group II), and 18 mothers treated for syphilis before delivery and their asymptomatic infants (group III). A fetal IgM response to T. pallidum antigens with apparent molecular masses of 72, 47, 45, 42, 37, 17, and 15 kDa was observed among sera of infants with congenital syphilis. Fractionation of sera into IgM and IgG components by high performance liquid chromatography confirmed that fetal IgM antibodies in every case were directed specifically against a 47-kDa antigen. Two asymptomatic infants from group II also showed serum IgM reactivities with the 47-kDa antigen, thereby appearing to confirm in utero infection. The combined data suggest that fetal serum IgM reactivity with the 47-kDa antigen of T. pallidum can be used as an important molecular marker for the diagnosis of congenital syphilis.     

Risk factors and consequences of congenital Chagas disease in Yacuiba, south Bolivia

Objective To determine the risk factors of congenital Chagas disease and the consequences of the disease in newborns. Methods Study of 2712 pregnant women and 2742 newborns in Yacuiba, south Bolivia. Chagas infection was determined serologically in mothers and parasitologically in newborns. Consequences of congenital Chagas disease were assessed clinically. Results The prevalence of Chagas disease in pregnant women was 42.2%. Congenital transmission was estimated at 6% of infected mothers leading to an incidence rate of 2.6% among newborns. Main risk factors of congenital transmission were mothers’ seropositivity and maternal Trypanosoma cruzi parasitaemia. Parity was higher in infected than in non‐infected mothers, but it was not associated with the risk of congenital transmission. The rate of congenital infection was significantly higher in newborns from multiple pregnancies than in singletons. However, we did not observe statistically significant consequences of Chagas disease in newborns from single pregnancies or among twins. Conclusions The main risk factors for congenital transmission were infection and parasitaemia of mothers. Consequences of the disease seemed mild in newborns from single pregnancies and perhaps more important in multiple births.     

Maternal Trypanosoma cruzi infection, pregnancy outcome, morbidity, and mortality of congenitally infected and non-infected newborns in Bolivia

This work compares the results of two epidemiologic and clinical surveys on the consequences of maternal chronic Trypanosoma cruzi infection. They were conducted in 1992-1994 and 1999-2001 in the same maternity clinic in Bolivia, a country highly endemic for infection with this parasite. In both surveys, the materno-fetal transmission of parasites occurred in 5-6% of the infected mothers. Maternal chronic T. cruzi infection had no effect on pregnancy outcome and health of newborns when there was no materno-fetal transmission of parasites. Comparisons between the older and the more recent surveys highlighted significant reductions in frequencies of symptomatic cases (from 54% to 45%), Apgar scores < 7, and low birth weights and prematurity (from 32-50% to 6-16%) among congenitally infected babies. Neonatal mortality related to congenital Chagas disease also decreased from 13% to 2% in the interval between both studies. These results suggest that the decrease in poverty that has occurred in Bolivia between both surveys might have contributed to reduce the morbidity and mortality, but not the transmission rate of T. cruzi congenital infection, which remains a serious public health problem in this country.     

Temporal changes in the humoral immune response of cattle during experimental infections with Onchocerca lienalis

In order to gain insights into the immune response in onchocerciasis during early infection, laboratory-reared calves were infected with 1000 Onchocerca lienalis infective larvae and examined serologically over a period of 508 days. Levels of serum antibodies measured by ELISA against adult worm extract revealed a multiphasic response, characterized by a broadly similar profile of peaks in individual animals arising at 15–30, 79 and >266 days after infection. Timings of these changes in responsiveness closely mirrored parasite development, coinciding with larval moults and with the onset of a patent infection. The levels of individual antibody isotypes directed against parasite antigens was strongly skewed. The dominant response was of IgG1, although limited reactivities were also found for IgG2 and IgM: No parasite-specific IgA antibodies were detected. Immuno-blots of adult worms extracts revealed a pattern of antigen recognition over time that matched the results obtained by ELISA. Again, the IgGl response was strongest, although certain lgG2 and IgM specificities were well represented. In general, there was a steady increase in the number of individual antigens recognized as the infection progressed, with a striking expansion of antibody specificities from day 79 following the fourth larval moult. Antibodies to a 16kDa component were a prominent feature of the response following development of a patent infection. These data reveal the strong influence of parasite biology on the development of the immune response in onchocerciasis.     

Association between measles antibodies in vaccinated and naturally infected mothers with protective antibodies and the occurrence of measles in their children:A cross-sectional study in the Bavi district of Hanoi

Objective:To determine the concentration and rate of decay of maternal IgG antibodies against measles prevalence in infants of vaccinated or naturally infected mothers and study initial measles immunization occurs in nine-month-old children.Methods:In total,401 pregnant women and the same number of their subsequent newborns were recruited in the Bavi district of Hanoi in 2016-2017;they were divided into two groups:Older women(born before 1985,n=201)and younger women(born after 1990,n=200).Samples were collected at five time-points;week 36 of pregnancy,birth(cord),and 3,6,and 9 months after birth.Measles-specific IgG antibody levels were recorded.Results:In total,77.06% of the 401 pregnant women were seropositive for measles-specific IgG antibodies.A significantly greater proportion of mothers aged 30 and older(88.06%)and their newborn(93.53%)were seropositive compared to the mothers aged 25 and younger(66.00%),and their newborn(72.00%)(P0.001).The infants of older mothers had significantly higher geometric mean titres(GMT)of measles IgG antibodies than the infants of younger mothers(P0.001)at all time-points of the study period.The proportion of measles IgG antibodies together with GMT decreased from 82.97%(506.96)at the age of three months to 23.19%(45.22)at the age of nine months.Conclusions:This study provides a profile of maternal antibodies against measles in Vietnamese infants and investigates the early susceptibility to measles in both the infants of vaccinated mothers and mothers with naturally acquired immunity.These data suggest that determining the appropriate age for measles vaccination is paramount for the elimination of measles in Vietnam.     

A survey of congenital Chagas' disease, carried out at three health institutions in São Paulo City, Brazil

The congenital transmission of Chagas' disease was evaluated in 57 pregnant women with Chagas' disease and their 58 offspring. The patients were selected from three Health Institutions in S?o Paulo City. The maternal clinical forms of Chagas' disease were: indeterminate (47.4%), cardiac (43.8%) and digestive (8.8%); 55 were born in endemic areas and two in S?o Paulo City. The transmission of Chagas' disease at fetal level was confirmed in three (5.17%) of the 58 cases studied and one probably case of congenital Chagas' disease. Two infected infants were born to chagasic women with HIV infection and were diagnosed by parasitological assays (microhematocrit, quantitative buffy coat-QBC or artificial xenodiagnosis). In both cases the placenta revealed T. cruzi and HIV p24 antigens detected by immunohistochemistry. In one case, a 14-week old abortus, the diagnosis of congenital T. cruzi infection was confirmed by immunohistochemistry. The other probable infection, a 30-week old stillborn, the parasites were found in the placenta and umbilical cord. The Western blot method using trypomastigote excreted/secreted antigens of T. cruzi (TESA) was positive for IgG antibodies in 54/55 newborns and for IgM in 1/55 newborns. One of the two newborns with circulating parasites had no detectable IgG or IgM antibodies. The assessment of IgG antibodies in the sera of pregnant women and their newborns was performed by ELISA using two different T. cruzi antigens: an alkaline extract of epimastigotes (EAE) and trypomastigote excreted/secreted antigens (TESA). The analysis showed a linear correlation between maternal and newborn IgG antibody titers at birth.     

In patients on prolonged HAART,a significant pool of HIV infected CD4 T cells are HIV-specific

OBJECTIVES: To examine the antigen specificities of HIV reservoir CD4 T cells in patients on prolonged and effective highly active antiretroviral therapy (HAART). DESIGN: Five HIV-infected patients, who were highly adherent to antiretroviral treatment, were selected on the basis of long-term undetectable plasma viral RNA on unmodified HAART. To investigate the antigen specificities of infected memory CD4 T cells, we examined the capacity of recall antigens, including HIV antigens, to induce virus production by peripheral blood mononuclear cells (PBMC). METHODS: To quantify CD4 T cells infected by replication-competent virus, and to determine their antigen specificities, we used a limiting dilution-based culture assay. CD8 T cell-depleted PBMC at several cell densities were activated by using Tuberculin purified protein derivative, cytomegalovirus, or HIV-1 p24 with and without HIV-1 Nef. RESULTS: We found that the pool of infected CD4 T cells includes HIV-specific cells with apparent frequencies between 5- and 100-fold higher than those of the common specificities for cytomegalovirus or Tuberculin. CONCLUSION: Our findings suggest that a significant proportion of replication-competent HIV-infected CD4 T cells in these patients are memory cells directed against HIV determinants. This may provide a rationale for the therapeutic use of recombinant HIV antigens to reduce the pool of HIV-reservoir cells.     

Cross–reactivity of antibody against an epitope of the Plasmodium falciparum second merozoite surface antigen

Monoclonal antibodies directed against the 51 kD merozoite surface antigen of Plasmodium falciparum also bind to other antigens within the infected cell. The sizes of these cross-reacting antigens have been characterized. Immunofluorescence due to the reaction of one of the monoclonal antibodies with these cross-reacting antigens was localized in the intra-erythrocytic parasite and in granules in the infected red cell cytoplasm. This immunofluorescence could be distinguished from the merozoite surface antigen in parasite lines with a variant serotype of the merozoite surface antigen which fails to react with the monoclonal antibodies. It was found that the in-vitro growth inhibition caused by the presence of one of the monoclonal antibodies, 8G10/48, was dependent on the expression of the corresponding serotype of merozoite surface antigen, a finding consistent with the inhibitory effect of this antibody being primarily directed against the merozoite surface antigen and not the cross-reacting antigens. Analysis of the frequency at which epitopes occur suggests that such cross-reacting proteins will be commonly seen in malaria, without the need to postulate a selective advantage for such cross-reacting specificities.     

Acquisition of antibody isotypes against Plasmodium falciparum blood stage antigens in a birth cohort

Information on the period during which infants lose their maternally derived antibodies to malaria and begin to acquire naturally their own immune responses against parasite antigens is crucial for understanding when malaria vaccines may be best administered. This study investigated the rates of decline and acquisition of serum antibody isotypes IgG1, IgG2, IgG3, IgG4, IgM and IgA to Plasmodium falciparum antigens apical membrane antigen (AMA1), merozoite surface proteins (MSP1‐19, MSP2 and MSP3) in a birth cohort of 53 children living in an urban area in the Gambia, followed over the first 3 years of life (sampled at birth, 4, 9, 18 and 36 months). Antigen‐specific maternally transferred antibody isotypes of all IgG subclasses were detected at birth and were almost totally depleted by 4 months of age. Acquisition of specific antibody isotypes to the antigens began with IgM, followed by IgG1 and IgA. Against the MSP2 antigen, IgG1 but not IgG3 responses were observed in the children, in contrast with the maternally derived antibodies to this antigen that were mostly IgG3. This confirms that IgG subclass responses to MSP2 are strongly dependent on age or previous malaria experience, polarized towards IgG1 early in life and to IgG3 in older exposed individuals.     

Detectable Trypanosoma cruzi parasitemia during pregnancy and delivery as a risk factor for congenital Chagas disease

Vector control has led to a drastic decrease in the prevalence of acquired Chagas disease in Latin America, thus redirecting attention to congenital Chagas disease. We report results of a longitudinal study of 359 pregnant women in Yacuiba in southern Bolivia, of whom 147 (40.9%) were infected with Trypanosoma cruzi, to evaluate the relationship between the patency period of the parasitemia and the risk of congenital infection. Maternal infection was assessed by using T. cruzi-specific serologic tests, and parasitemia in mothers and newborns was diagnosed by using microscopic examination of blood in heparinized microhematocrit tubes. Parasitemia was present in 28.6% of the infected women. Its prevalence increased during the third trimester, then decreased at delivery. The likelihood of congenital infection was significantly correlated with the parasite density in the mother's blood. The risk of transmission increased during the third trimester of pregnancy and could explain premature births or low-weight newborns for infected mothers.     

Congenital Chagas disease of second generation in Santiago, Chile. Report of two cases

Congenital Chagas disease (CChD) has been reported in different countries, mostly in Latin America. In 1987 a fatal case of CChD of second generation (CChDSG) was published. Within a period of six months--1989-1990--two cases of CChDSG were diagnosed and studied in the city of Santiago. Two premature newborns, sons of two sisters, with moderate liver and spleen enlargement, were found to have positive serology for Chagas disease and xenodiagnoses. The mothers, urban residents all their lives, without antecedents of triatomine bugs contact or blood transfusions, showed positive serology and xenodiagnoses. Their mother (grandmother of the infants), lived 20 years in a Northern rural Chagas disease endemic locality, in a triatomine infested house. Afterwards, she moved to Santiago, where she married and has resided up to now. Serology and xenodiagnoses were also positive. All the Trypanosoma cruzi infected individuals were successfully treated with nifurtimox.     

Antibody responses in schistosomiasis haematobium in Somalia. Relation to age and infection intensity

Antibody responses in schistosomiasis haematobium were studied in relation to age and infection intensity in Somalia. The area is highly endemic for Schistosoma haematobium but free of S. mansoni. Antibodies of the IgG class against particulate antigens of S. mansoni adult worms were investigated by immunofluorescence (gut and somatic associated antigens) and against soluble egg and adult worm antigens by ELISA. Total IgE levels were examined by Pharmacia IgE RIA, and specific IgE against soluble adult worm antigen by enzyme immunoassay. The IgG antibody response showed a characteristic pattern with highest reactivity against both gut associated and soluble egg antigens in the age group 10-14 years, when both prevalence and intensity of the infection were highest. Reactivity against somatic associated antigen was also high in this age group, but it increased slightly and remained at high level in the older ages. It is thought that such antigen is exposed mainly after the death of the parasite and that the antigenic stimulation may remain throughout most of the life of infected individuals. On the other hand, the IgG antibody reactivity against soluble adult worm antigen was low during childhood, but it increased significantly with age. It is suggested that repeated booster effects are needed for more potent response against these antigenic components. The finding of high levels of total IgE already in the youngest age groups, together with low specific IgE response, indicates that mainly other antigens are involved in the IgE production. The specific IgE response against soluble adult worm antigen was low but increased significantly with age.     

Comparison of Trypanosoma cruzi lineages and levels of parasitic DNA in infected mothers and their newborns

To better understand the factors involved in maternal-fetal transmission of Trypanosoma cruzi, we compared DNA levels-obtained by use of quantitative real-time PCR and parasitic genotypes determined by PCR amplification followed by hybridization-in Bolivian mothers and their congenitally infected newborns. Mothers and their neonates displayed markedly different parasitic DNA levels, as most maternal estimated parasitemias (> 90%) were < 10 parasites/mL, whereas those of 76% of their newborns were > 1,000 parasites/mL. Comparison of T. cruzi TcII sublineages infecting mothers and newborns showed identity, without evidence of mixed infection in mothers or neonates. Analysis of minor variants of TcIId-genotyped parasites using sequence class probes hybridizing with hypervariable domains of kDNA minicircles showed discrepancies in half of mother/newborn pairs.