Establishing reference ranges for lymphocyte proliferation responses to phytohemagglutinin in patients with T cell dysfunction

Introduction: The evaluation of lymphocyte proliferation responses is a critical component of the clinical work up for patients with suspected immunodeficiencies. Those with severe combined immunodeficiency (SCID) have consistently low to absent responses (stimulation index, SI) to the mitogen phytohemagglutinin (PHA). However, patients with combined immunodeficiency (CID) have more varied proliferative responses, and are open to a wide range of interpretations. Aims: To establish lymphocyte proliferation response reference ranges for patients with T cell defects, especially those with CID as well as healthy controls. Methods: Data was collected retrospectively from charts of patients with a diagnosis of SCID (n = 39), CID (n = 52), or from healthy controls (n = 440). Reference percentiles were calculated using the 95% of the distribution of the test results. Results: The reference ranges for the control group ranged from 134 to 2220.5, whereas those with CID were distributed between 0.81 and 169.1. Patients with typical SCID had profound low proliferative responses, with SI <5. Conclusion: Our results highlight the variability of lymphocyte proliferation responses to PHA in patients with CID as well as healthy controls. These reference ranges will assist with the critical interpretation of assay results, particularly when values fall on the extreme end of the range. Statement of novelty: We provide reference ranges for lymphocyte proliferation responses to PHA from patients with CID and healthy controls.

Micronutrient supplementation and T cell-mediated immune responses in patients with tuberculosis in Tanzania

SUMMARYLimited studies exist regarding whether incorporating micronutrient supplements during tuberculosis (TB) treatment may improve cell-mediated immune response. We examined the effect of micronutrient supplementation on lymphocyte proliferation response to mycobacteria or T-cell mitogens in a randomized trial conducted on 423 patients with pulmonary TB. Eligible participants were randomly assigned to receive a daily dose of micronutrients (vitamins A, B-complex, C, E, and selenium) or placebo at the time of initiation of TB treatment. We found no overall effect of micronutrient supplements on lymphocyte proliferative responses to phytohaemagglutinin or purified protein derivatives in HIV-negative and HIV-positive TB patients. Of HIV-negative TB patients, the micronutrient group tended to show higher proliferative responses to concanavalin A than the placebo group, although the clinical relevance of this finding is not readily notable. The role of nutritional intervention in this vulnerable population remains an important area of future research.

Characterization of a Novel Inactivated Salmonella enterica Serovar Enteritidis Vaccine Candidate Generated Using a Modified cI857/λ PR/GeneEExpression System

ABSTRACTA new strategy to develop an effective vaccine is essential to control food-borneSalmonella entericaserovar Enteritidis infections. Bacterial ghosts (BGs), which are nonliving, Gram-negative bacterial cell envelopes, are generated by expulsion of the cytoplasmic contents from bacterial cells through controlled expression using the modified cI857/λ PR/geneEexpression system. In the present study, the pJHL99 lysis plasmid carrying the mutated lambda pR37-cI857 repressor and PhiX174 lysis geneEwas constructed and transformed inS. Enteritidis to produce a BG. Temperature induction of the lysis gene cassette at 42°C revealed quantitative killing ofS. Enteritidis. TheS. Enteritidis ghost was characterized using scanning and transmission electron microscopy to visualize the transmembrane tunnel structure and loss of cytoplasmic materials, respectively. The efficacy of the BG as a vaccine candidate was evaluated in a chicken model using 60 10-day-old chickens, which were divided into four groups (n= 15), A, B, C, and D. Group A was designated as the nonimmunized control group, whereas the birds in groups B, C, and D were immunized via the intramuscular, subcutaneous, and oral routes, respectively. The chickens from all immunized groups showed significant increases in plasma IgG and intestinal secretory IgA levels. The lymphocyte proliferation response and CD3+CD4+and CD3+CD8+T cell subpopulations were also significantly increased in all immunized groups. The data indicate that both humoral and cell-mediated immune responses are robustly stimulated. Based on an examination of the protection efficacy measured by observations of gross lesions in the organs and bacterial recovery, the candidate vaccine can provide efficient protection against virulent challenge.

In VitroImmune Competence of Buffaloes (Bubalus bubalis) of Different Production Potential: Effect of Heat Stress and Cortisol

Twelve healthy lactating Murrah buffaloes of similar parity (3rd) between 90 and 120 days of lactation, selected from the herd of National Dairy Research Institute (Karnal, India) and maintained at managemental practices as followed at the Institute they were included in this experiment. The animals were divided into two groups based on their production level in previous lactation. The average milk production level of group 1 and II was 9.3 and 6 lit/day, respectively. Blood was collected from these buffaloes on three occasions 10 days apart. The lymphocytes were separated and cultured in RPMI 1640 medium with PHA-P for 24 h at 37°C in a humidified CO2incubator (95% air and 5% CO2). The lymphocyte responsiveness was also evaluated in response to thein vivoheat stress andin vitrocortisol. Mitogen-induced stimulation index was not affected by production level (). Stimulation index was significantly reduced () in both the groups when cortisol was added at 2.0 ng level in the culture. However, in heat-stressed buffaloes stimulation index did not vary despite increasing levels of cortisol, thus indicating that lymphocyte may become cortisol resistant during periods of acute heat stress. The results showed that lymphocyte proliferation response can be effectively used to study buffalo cell-mediated immunityin vitro.

Impact of feed-borne mycotoxins on avian cell-mediated and humoral immune responses

Mycotoxins of economic importance in poultry production are mainly produced by Aspergillus, Penicillium and Fusarium fungi. The important mycotoxins in poultry production are aflatoxins, ochratoxins, trichothecenes, zearalenone and fumonisins. Mycotoxins exert their immunotoxic effects through various mechanisms which are manifested as reduced response of the immune system. Mycotoxin-induced immunosuppression in poultry may be manifested as decreased antibody production to antigens (e.g. sheep red blood cells) and impaired delayed hypersensitivity response (e.g. dinitrochlorobenzene), reduction in systemic bacterial clearance (e.g. Salmonella, Brucella, Listeria and Escherichia), lymphocyte proliferation (response to mitogens), macrophage phagocytotic ability, and alterations in CD4+/CD8+ ratio, immune organ weights (spleen, thymus and bursa of Fabricius), and histological changes (lymphocyte depletion, degeneration and necrosis). Mycotoxins, especially fumonisin B1 have been shown to down regulate proinflammatory cytokine levels including those of interferon (IFN)-γ, IFN-α, interleukin (IL)-1β, and IL-2 in broiler chickens. Fusarium mycotoxins exert part of their toxic effects by altering cytokine production in poultry. Mycotoxins adversely affect intestinal barrier functions by reducing the intestinal epithelial integrity and removing tight junction proteins. Apoptosis, increased colonisation of pathogenic microorganisms, cytotoxicity and oxidative stress, inhibition of protein synthesis and lipid peroxidation are characteristic of the toxic effects of mycotoxins on intestinal epithelium. These directly or indirectly affect host immune responses. Such immunotoxic effects of mycotoxins render poultry susceptible to many infectious diseases. The avian immune system is sensitive to most mycotoxins. Both cell-mediated and humoral immunity may be adversely affected after feeding mycotoxins to poultry. The avian immune system may be more sensitive to naturally contaminated feedstuffs because of the presence of multiple mycotoxins and the complex interactions between them which can cause severe adverse effects. Adverse effects of mycotoxins on the immune system reduce production and performance resulting in economic losses to poultry industries. Caution must be exercised while feeding grains contaminated with mycotoxins.