Neonatal tetanus despite protective serum antitoxin concentration

Using the ELISA technique to estimate serum antibodies against tetanus toxin, seven neonates with clinical tetanus were found to have antibody levels 4-13 times higher than the presumed minimum protective level of 0.01 IU/ml. All but one of their mothers had been vaccinated with tetanus toxoid in pregnancy. In two other neonates, whose mothers had received multiple booster doses of toxoid during pregnancy, the anti-toxin concentrations were 100- and 400-times the presumed protective level. Therefore the toxin dose may overwhelm the pre-existing anti-toxin level and produce disease. Furthermore, multiple booster injections of tetanus toxoid may not only enhance serum anti-toxin titres, but could also lead to an ineffective immune response.     

Improving tetanus prevention in the Krasnodar area taking the current epidemic situation into account]

An increase of percentage of elderly persons among those who fell ill with tetanus (70%) was noted against a sharp reduction of tetanus incidence under the effect of mass active immunization against this disease. A study of immunity in older and elderly individuals showed the percentage of immune persons among them to be rather low (48.8--55.6%). Due to difficulty of elderly individuals embracement by vaccination the authors suggest a single immunization scheme with a double toxoid dose (20 BU) followed by revaccination with 10 BU in one year. A total of 21 472 persons were placed under observation. The suggested immunization scheme was harmless, promoted stimulation of antitetanus immunity in persons vaccinated earlier; as to unvaccinated persons--it created a favourable immunological preparedness for revaccination, permitting to do without any antitetanus serum in case of trauma.     

In vitro studies of the rabbit immune system. VI. Rabbit anti-mouse cytotoxic T effector cells are inhibited by anti-rabbit T cell serum in the absence of complement.

Xenogeneic rabbit anti-mouse cell-mediated cytotoxic activity could be generated by culturing lymphoid cells from mesenteric lymph nodes (MLN), spleen, or peripheral blood of rabbits primed 2 to 8 weeks earlier with mouse tumor or spleen cells. MLN cells, which provided the best source of activity after being cultured with 5 to 10 X 10(6) mitomycin C-treated mouse spleen cells for 4 to 6 days, produced 30 to 90% specific isotope release after 4 to 7 hr incubation with 15Cr-labeled tumor target cells. Xenogeneic cytotoxic activity was primarily H-2 specific and could not be blocked by immune complexes but was abrogated by treatment with goat anti-rabbit thymocyte serum plus complement (ATS + C) before or after culture. Therefore, the activity appeared to be mediated by cytotoxic T lymphocytes (CTL). Furthermore, ATS without C abrogated cytotoxic activity when included in the CTL assay at concentrations of 5 to 15 microliter/10(7) effector cells. The inhibitory activity of ATS was directed to the rabbit effector population and could be absorbed completely by rabbit thymocytes. Antisera to mouse T cells with comparable cytolytic activity in the presence of C did not inhibit murine allogeneic CTL.     

Microtissue Culture Test for the Titration of Low Concentrations of Diphtheria Antitoxin in Minimal Amounts of Human Sera

A microtissue culture method for the assay of low concentrations of diphtheria antitoxin in human sera has been developed, using a monkey kidney cell (VERO) culture technique. Results obtained with sera from nonvaccinated children and with immune sera from children vaccinated with three and four injections of diphtheria pertussis tetanus vaccine were in agreement with antitoxin levels considered necessary to denote immunity to diphtheria. The use of microplates and organic buffer for culturing the animal cells improved the stability of the tissue culture system. The described method is sensitive, economical, and applicable for the titration of antitoxin in human sera particularly from infants and children from whom a minimum amount of serum is available.     

Regualtion of the immune response to tumor antigens. I. Immunosuppressor cells in tumor-bearing hosts.

A/Jax mice were rendered immune to the syngeneic and transplantable methylcholanthrene-induced Sarcoma 1509a by the surgical removal of the tumor 7 days after implantation; subsequent injection i.v. transfer of 10(7) to 10(8) washed thymus or spleen cells of tumor-bearing animals (TBA) to immune animals significantly inhibited the rejection of the tumor; this suppressive effect was entirely abolished by the treatment of these lymphocytes with anti-theta serum or anti-thymocyte serum (ATS) and complement before adoptive transfer. On the other hand, an equal number of thymus or spleen cells of normal animals or of animals bearing an unrelated tumor had no suppressive effect. Treatment of normal syngeneic animals with ATS after tumor cell inoculation or splenectomy of TBA resulted in the suppression of the tumor growth. The serum of TBA had no effect on tumor growth in immune syngeneic mice. Together these results suggest that TBA possess immunosuppressor T cells regulating negatively their immune response to the tumor.     

Trypanosoma cruzi: effects of anti-thymocyte serum in mice and neonatal thymectomy in rats

CF₁ mice were given eight injections of normal rabbit serum (NRS), Hanks' balanced salt solution (HBSS), or rabbit anti-mouse thymocyte serum (ATS) beginning 3 days prior to and at 3-day intervals subsequent to intraperitoneal (ip) inoculation with 5 × 10⁴ trypomastigotes of a Brazil strain of Trypanosoma cruzi. Markedly enhanced parasitemia, increased numbers of tissue stages (amastigotes), and higher mortality occurred in ATS-treated mice as compared to NRS- or HBSS-treated controls. Administration of three injections of ATS at 3-day intervals during the latter stages of acute Chagas' disease, i.e., when numbers of parasites were declining, resulted in a transitory relapse (increase in numbers) of blood and tissue parasites. No relapse occurred in mice when ATS was administered at 3-day intervals over a period of 15 days during the subacute stage of the disease, i.e., after parasites had disappeared from the blood.Parasitemia and mortality were enhanced in neonatally thymectomized rats when compared to that observed in sham-operated and unoperated control rats following ip injection of 2 × 10⁵ trypomastigotes of T. cruzi. Serum obtained from thymectomized and control rats 5 weeks after inoculation with T. cruzi at a time when the blood of all animals had become microscopically negative for parasites were equally protective in passive transfer experiments, while serum from uninfected controls gave no protection.Gamma globulin levels significantly increased in thymectomized as well as intact rats by the third to fourth week of infection with T. cruzi, reached maximum concentrations in 5–6 wk, and remained elevated significantly at the twelfth week post infection as compared with uninfected controls. No significant changes occurred in total serum proteins or α and β fractions of any group, infected or uninfected.Total circulating leukocytes, especially lymphocytes, were diminished in mice and rats subjected to treatment with ATS or neonatal thymectomy.These data clearly indicate that neonatal thymectomy of rats and ATS treatment of mice suppress the acquired immune response to T. cruzi. Further, passive transfer experiments in rats confirm the protective role of circulating antibody in acquired immunity to Chagas' disease.     

Tetanus antibody titers and duration of immunity to clinical tetanus infections in free-ranging rhesus monkeys (Macaca mulatta)

Prior to 1985 tetanus was a major cause of mortality in the free-ranging colony of rhesus monkeys on Cayo Santiago, accounting for almost a quarter of annual deaths. In 1985 and 1986 all animals (except infants) received primary and booster doses, respectively, of tetanus toxoid. In subsequent years primary immunizations were given to all yearlings, and boosters were administered to all 2-year-old animals during the annual capture of the colony. The main objectives of the tetanus immunization program were to reduce the pain and suffering caused by tetanus infections and to decrease mortality in the colony. Other objectives were to evaluate the efficacy of the two-dose tetanus toxoid immunization protocol and to determine whether additional boosters might be required to provide adequate long-term protection against tetanus infections. The immediate effect of the mass immunization program was the elimination of clinical tetanus infections in the population and a 42.2% reduction in the overall mortality rate. Since the immunization program began, no cases of tetanus have been observed in the colony, except in two unimmunized infants, and it has not been necessary to give tertiary injections of tetanus toxoid to maintain protection against infection. A sample collected in 2004 of the original cohort of monkeys immunized in 1985 and 1986 showed that 93.3% (14/15) had protective tetanus antibody titers (>0.01 IU/ml) at the ages of 20-23 years, which is close to the life expectancy of the Cayo Santiago rhesus macaques. Two intramuscular doses of tetanus toxoid provided long-term, if not lifelong, protection against tetanus for rhesus monkeys living in a tropical clime where tetanus is enzootic and the risk of infection is great.     

Complement dependence of antibody-induced mesangial cell injury in the rat

Intravenous administration of rabbit anti-rat thymocyte serum (ATS) reactive with Thy-1-like antigens present on rat mesangial cells induces almost immediate (1-hr) mesangial cell injury in rats followed by sequential mesangiolytic and mesangial-proliferative/infiltrative lesions. To determine the role of complement in these ATS-induced glomerular lesions, ATS was given to Lewis rats that had been depleted of C3 by cobra venom factor (CVF). CVF treatment prevented the degenerative changes in mesangial cells and accumulation of even the few polymorphonuclear leukocytes (PMN) seen in the glomeruli (2.67 PMN/glomerulus) 1 hr after ATS-treatment in rats not given CVF. In addition, CVF prevented the mesangiolysis and mesangial hypercellularity seen at day 4. Rat C3 and late complement components identified in the mesangial of ATS-treated rats in close association with the deposition of rabbit immunoglobulin G was also absent as a result of CVF treatment. CVF treatment did not affect binding of ATS to glomeruli as studied by immunofluorescence or paired label radioisotope techniques. The depletion of leukocytes and/or PMN by irradiation or treatment with anti-I-MN serum had no effect on the induction of the acute mesangial cell damage or the mesangiolytic lesion. Irradiation did diminish the 4-day proliferative/infiltrative lesion. Complement depletion normalized the ATS-induced increase in mesangial uptake of heat-aggregated human gamma-globulin (655.0 +/- 35.2 micrograms in ATS-treated vs 20.3 +/- 2.9 micrograms/5 X 10(4) glomeruli in ATS plus CVF-treated rats; mean +/- SEM). Small immune deposits present in the mesangial areas of kidneys 4 to 5 days after CVF treatment represented CVF-anti-CVF antibody-C3 complexes. The model of mesangial cell damage induced by ATS in the rat is complement-dependent and may relate, at least in part, to complement-mediated mesangial cell lysis.     

Epidemiological and immunological features of tetanus under conditions of long-term planned vaccine prophylaxis

The introduction of mass immunization against tetanus has resulted in the decrease of morbidity rate (5.2 times), the leveling of morbidity rate among the urban and rural population and among males and females, though no essential effect on the seasonal distribution of tetanus morbidity has been observed. Persons over 50 years of age (housewives and pensioners) have become the main groups of risk at the post-immunization period. Mass immunization against tetanus over a period of many years has ensured the existence of a sufficient immune stratum (89.9 +/- 3.0% to 100 +/- 3.0%) and a sufficient level of antitoxic immunity (means geom equal to 6.72-9.6 I.U./ml) among children. Among adults, the proportion of persons protected from tetanus decreases in older age groups from 82.1 +/- 1.3% in persons aged 31-40 years to 22.1 +/- 2.0% in persons over 60 years. The observed differences between the coverage of the population with immunization and the proportion of persons having protective titers of tetanus antibodies require constant control of the intensity of immunity and its correction with regard to its initial level, especially in persons of older age groups.     

Sublingual immunization with an engineered Bacillus subtilis strain expressing tetanus toxin fragment C induces systemic and mucosal immune responses in piglets

Sublingual (SL) and intranasal (IN) administration of a Bacillus subtilis-based tetanus vaccine was tested in piglets, which more closely mimic the human immune system than mice. Piglets were immunized by the SL, IN or oral routes with vaccine expressing tetanus toxin fragment C, or commercial tetanus vaccine given by intramuscular injection as a control. Tetanus toxoid specific ELISA and passive neutralization tests were used to measure IgG and IgA levels in serum and mucosal secretions, and assess protective serum antibodies, respectively. The nature of the immune response was explored by MHC Class II, TGF-β1 expression, and ELISA assays for multiple cytokines. SL or IN immunization of piglets induced neutralizing tetanus toxoid specific serum antibody and local salivary and vaginal IgA responses. Standard tetanus vaccine resulted in systemic antibodies, whereas oral administration of the Bacillus-based vaccine was ineffective. Further analyses indicated a balanced Th1/Th2 response to SL or IN immunization. CONCLUSION: This study demonstrates for the first time that SL or IN administration is effective for inducing both systemic and mucosal responses in a piglet model, indicating that SL or IN delivery of a B. subtilis-based tetanus vaccine can be a simple, non-invasive, low cost strategy to induce immunity to tetanus.     

人淋巴细胞的体外抗体诱发及其在杂交瘤中的应用

Sources of immunized lymphocytes constitute one of the main obstacles in the production of human monoclonal antibodies. We tried to get them through in vitro immunization. Cells from excised tonsils or trauma spleens were used for the induction of antibody responses in vitro. Antibodies to different antigens including sheep red blood cells, ovalbumin, tetanus toxoid, and hepatitis B surface antigens were induced in 7-14 days' cultures. Taking tetanus toxoid as antigen, we analysed the various factors required for antibody induction with statistics analysis, which included cell separation method, T cell conditioned medium, antigen dosage, serum content, and concentration of mitogen PWM and LPS. The results showed: (1) The cell separation method influenced the antibody production significantly in comparison with other factors. It signified that immune cells' combination was the most influential factor. (2) Serum also constituted quite important influencing factor especially in the later period of culture. However, it did not make much differences if it attended 10% or so. The antigens and mitogens tended to be used at low concentration. (3) Due to the significant variation among individuals and among different antigens, it is suggested to set up the culture system with some flexibility so as to adapt to the variation in cells and antigens from different sources. The present culture system we use includes nylon wall column separation of cells, suitable range of antigens (three doses instead of one), and either 10% T cell conditioned medium or a mixture of 1 microgram PWM/ ml + 0.1 microgram LPS/ml. The human B lymphocytes stimulated in vitro with tetanus toxoid were used for the construction of human hybridomas.     

The effect of specific anti-rat thymocyte serum on cell-mediated tumor immunity and other lymphocyte functions

A burro anti-rat thymocyte serum (ATS) was rendered specific for thymus-derived lymphocytes (T cells) by repeated absorptions with Murphy-Sturm lymphoma cells. All of the cells in this lymphoma cell line were shown to have surface immunoglobulin and it was presumed to be bone marrow-derived. Treatment of lymphocytes with ATS eliminated stimulation by phytohemagglutinin and concanavalin A, but did not reduce the proportion of cells with surface immunoglobulin or complement receptor. ATS also did not decrease the number of antibody-producing spleen cells from rats immunized against sheep erythrocytes. The ATS was used to determine the nature of the effector cells in the cell-mediated cytotoxicity of immune W/Fu rat spleen cells against a syngeneic Gross virus-induced lymphoma. This cytotoxic response was consistently inhibited, indicating that T cells were needed for reactivity.     

Role of T- and B-lymphocytes in the heterogeneity of human cell-mediated reactions to bacterial antigens]

Leucocytes from 30 patients with allergy to tuberculin and bacterial antigens were treated with antithymus (ATS) and anti-immune globulin (AIGS) sera. The leucocyte migration inhibition test (LMIT) was performed with these antigens. ATS abolished the LMIT induced by tuberculin and sometimes by bacterial antigens (staphylococcal, streptococcal etc.). AIGS frequently abolished the LMIT induced by bacterial antigens, but not by tuberculin. In some cases the treatment with any serum abolished the LMIT induced by the antigens, or, on the contrary, it was abolished only by a successive treatment with both sera. The lymphocyte types (T or B) determining the secondary immune response to the same antigen are different in various patients, as well as they differ in the same patients in relation to diverse antigens. Five types of lymphocyte - antigen interrelation in the LMIT have been distinguished.     

In vitro studies of the rabbit immune system. IV. Differential mitogen responses of isolated T and B cells.

The mitogenic responses of separated rabbit lymphocyte populations functionally analogous to mouse T and B cells have been tested in vitro. Purified T cells were prepared by passage over nylon wool (NW) and purified B cells prepared by treatment with antithymocyte serum and complement (ATS + C). ATS + C kills 70% of peripheral blood lymphocytes (PBL's) and 50% of the spleen cells while passage over NW yields 40% of the applied PBL's and 5–23% of the applied spleen cells. NW-purified T cells from the spleen or PBL's respond fully to concanavalin A (Con A) but have a reduced response to phytohemaglutinin (PHA) and little or no response to goat anti-rabbit immunoglobulin (anti-Ig). PBL's that survive ATS + C (B cells) are stimulated by anti-Ig but not by Con A or PHA. B cells purified from spleen do not respond to Con A or PHA but will respond to anti-Ig under appropriate conditions. A full spleen B-cell response to anti-Ig required removal of Ig produced by the cultures that blocked anti-Ig stimulation. It is concluded that, for rabbit lymphocytes, Con A and PHA are primarily T-cell mitogens and that anti-Ig is primarily a B-cell mitogen. However, the mitogen response of unfractionated PBL or spleen cell populations indicates an overlap in reactivity. This could be due to cells sharing T and B properties, alteration of cell populations by the fractionation procedures used, or recruitment of one population in the presence of a mitogenic response of the other population.     

贵州省典型植烟生态区域根际土壤微生物群落多样性

土壤微生物是土壤的重要组成部分,它对土壤肥力的形成和植物营养的转化起着积极的作用。采用16S V4区高通量测序等方法对贵州省不同生态区域典型植烟土壤根际微生物群落进行分析研究,从根际土壤微生物群落特征上诠释不同生态区域典型土壤根际微生物群落的信息差异。结果表明:贵州省不同生态区域弱酸和酸性土壤占70.06%,有机碳平均2.24%,有机碳含量总体偏高,全氮与有机碳含量具有一定相关性;根际土壤微生物的OTUs数量差异明显,呈现总体是西部高于东部,南部高于北部;微生物属水平物种丰度差异显示,土壤类型对微生物群落丰度影响显著,而在土壤功能微生物上黔北区域的变形菌门(Proteobacteria)和放线菌门(Actinobacteria)两个门的微生物相对丰度最高,而在黔中中部区域变形菌门的丰度显著高于放线菌门的丰度。黔西南和黔中中部区域土壤微生物属水平优势种群显著高于其他地区,种群平衡度不高,具有土传病害发生的潜在危险。通过贵州省不同生态区域土壤微生物根际微生物信息分析与研究,为后续土壤生物修复奠定了良好的研究基础。     

In vitro studies of the rabbit immune system: III. T-cell requirement for initiation but not maintenance of anti-sheep erythrocyte plaque-forming cells

The in vitro anti-sheep erythrocyte plaque-forming-cell (anti-SRBC PFC) response of primed rabbit spleen cells can be abrogated by prior treatment with anti-thymocyte serum plus complement (ATS + C). The direct addition of ATS without C to stimulated cultures also reduces the response 60–90%, if the antiserum is added before the initiation of antigen-stimulated PFC production. However, the PFC response becomes refractory to ATS inhibition by 48 hr. This loss of ATS sensitivity is not caused by T cells themselves becoming refractory to ATS for the following reasons: (i) ATS + C treatment or ATS addition at 48 hr causes cytotoxic effects comparable to similar treatments at 0 hr; (ii) The responsiveness of cells incubated 48 hr before the addition of SRBC remains sensitive to ATS inhibition at that time. The loss of ATS sensitivity is also not the result of soluble T-cell factors which have accumulated in the medium, because the number of PFC in stimulated cultures continues to increase after treating with ATS + C and reculturing in fresh medium at 48 hr. We conclude that T cells are required to initiate the production of antibody by B cells but are not required to maintain the later proliferative events.     

Identification of human peripheral T cell antigens.

A new type of differentiation antigens on human T cells was demonstrated by using a heterologous anti-human T cell serum (ATS). This type of antigen, referred to as human peripheral T cell antigen (HPTA), was found on peripheral T cells and medullary thymocytes, but not on cortical thymocytes and B cells. The percentage of ATS-reactive lymphocytes in human peripheral lymphoid organs was correlated with that of cells rosetting with sheep erythrocytes, but contrasted with the number of B cells defined by the presence of a complement (C) receptor or by rabbit anti-human B cell serum (ABS). ATS also reacted with T cells purified by nylon fiber column filtration but ABS did not. Chronic lymphocytic leukemia cells rosetted with either sheep erythrocytes or erythrocyte-antibody-complement complexes were lysed by ATS and ABS, respectively. Mitogenic responses of blood lymphocytes to phytohemagglutinin (PHA) and concanavalin-A (Con A) were abrogated by treating them with ATS and C, whereas ABS suppressed only their response to Con A. Although numerous thymus cells rosetted with SRBC, only 14% were reactive with ATS. Quantitative absorption studies demonstrated that HPTA content of the thymus cells was much lower than that of lymph node cells. Anatomical localization of ATS-reactive lymphocytes in human lymphoid organs studied by immunofluorescence indicated that they were present in the thymus-dependent paracortical areas of lymph node and in the medullary region of thymus. ABS, on the other hand, did not stain thymocytes but reacted selectively with the cells located in the lymphoid follicles of lymph node. These data, together with that from cell suspension studies, confirmed that HPTA were shared between medullary thymocytes and peripheral T cells.     

Progress in Tetanus Prophylaxis: The Advent of Human Antitoxin

Injections of tetanus antitoxin of animal origin frequently cause serious disability and sometimes death. Despite world-wide knowledge of these effects, millions of prophylactic injections of equine tetanus antitoxin are given annually, and it is continually proposed that the dosage be increased in order to obtain higher “protective” levels in the serum, a procedure which would increase the incidence and severity of reactions. Furthermore, equine antitoxin frequently fails to prevent tetanus.Tetanus antitoxin of human origin is available which carries no risk of complications and confers a higher degree of immunity more quickly than equine antitoxin. The cost of treating reactions to horse serum, together with the financial loss incurred by work-absence, far outweighs the cost of human antitoxin. In the author''s opinion, the use, in this country, of antitoxin of animal origin is no longer medically acceptable and may well prove legally indefensible.     

Antidiphtheria and antitetanus antitoxic immunity indices in the population of the Estonian SSR (an assessment of the effectiveness of a planned vaccine prophylaxis)]

In 1972-1974 and 1977 in the Estonian SSR children and adults were surveyed for the presence of antibodies against tetanus and diphtheria toxins (toxoids) by means of the passive hemagglutination test. The level of protection against tetanus was revealed to correspond to the proportion of child population covered by vaccination: in 1977, with 98.8% covered by vaccination, the level of protection among children aged 7 to 14 years and adolescents of 15-19 years exceeded 98%; with the increase of age (every 10-15 years) the level of protection against tetanus regularly decreased. This dynamics correlated with the existing terms of postvaccinal immunity and the epidemiological independence of tetanus as infectiion. The level of protection in child population against diphtheria in 1972-1974 and 1977 lagged behind the level of protection against tetanus and the coverage by vaccination. The diphtheria component of adsorbed DPT vaccine seemed to be unable to ensure the sufficient level and intensity of immunity under conditions of a sharply decreased risk of encounter with the infective agent. In persons aged 40 years and over the indices of immunity against diphtheria were higher than against tetanus. These indices resulted from diphtheria infection at the prevaccination period and could serve as an objective sign in following up the decrease of the process of diphtheria epidemics.     

Effect of the joint use of BCG and antithymocyte serum on the growth of a syngeneic tumor in mice

The growth of the syngeneic tumor Acatol in BALB/c mice was retarded if the animals were pretreated with BCG or antilymphocyte serum (ATS). Combined use of BCG and ATS led to a significantly more powerful retardation as compared to the effect produced by each factor alone. Using the adoptive transfer of splenocytes from treated mice it was shown that tumor growth suppression is effected by the cell types other than T lymphocytes and macrophages. It is probable that the effector cells within the given system are K cells and natural killer cells. The results attest to a possibility of search for a two-directional action on the immune system of the tumor host, which would stimulate antitumor effector cells and inhibit the activity of suppressors, particularly that of T suppressors.