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The physical and chemical properties of a hemagglutinin from whole-body homogenates of the cephalochordate, Branchiostoma lanceolatum, are described. The hemagglutinin is proteinaceous since it is precipitated by trichloroacetic acid and ammonium sulphate, and all activity is lost at 60 degrees C or by treating with proteases. Carbohydrate moieties are probably also present since activity is lost after incubation with sodium metaperiodate. Activity is stable over pH 6-10. The agglutinin does not require Ca++ or Mg++, and a reduced titer after treatment with 2-mercaptoethanol and urea suggests the presence of both disulphide bonds and noncovalent linkages. Haemagglutination inhibition experiments with 17 saccharides and glycoproteins failed to show clear-cut carbohydrate specificity, with only mucin and fetuin having any inhibitory effect, so that the binding may be complex. Finally, cross-adsorption experiments suggest that only one lectin is present. The function of this lectin, especially in an immunobiological context, remains speculative.

Treatment of Extatosoma haemocytes on a monolayer with fixed rabbit erythrocytes (RE) in the presence of Ca2+ results in ca. 11% plasmatocytes (PL) and 58% spreading granular (SG) cells rosetting. Pretreatment of haemocytes with lactose, D-galactose, or asialofetuin reduces rosetting, suggesting that the membrane-associated receptors involved may be lectins. The percentage of rosetting PL and SG cells is increased by a number of heterologous lectins (indicating the presence of D-galactose, mannose, and Glu Nac receptors on these haemocytes) and a D-galactose specific lectin isolated from the serum of the insect. The latter acts as a bridging molecule and requires Ca2+ to bind to specific receptors on both haemocytes and fixed RE.

The classical definition of immunity as the resistance of the body to disease, views the immune system in simple mechanistic terms. In this brief overview, possible reasons and consequences of the presence of an effective immune system across a range of invertebrate and vertebrate animals are considered. Topics discussed include the forces favouring the development of immunity, such as the acquisition of the colonial habit and terrestrial mode of life by many primitive animals, the constant threats of microbial and macrobial invasion, and the need to eliminate somatic mutations. The consequences of immunity in terms of the development of autoimmunity and hypersensitivity reactions, as well as the interaction of the immune system with the brain, the neuroendocrine organs and environmental factors, are also examined. Finally, the possibilities that the delicate balance maintained between many parasites and the immune systems of their hosts results in the spread of disease, and may also determine the maintenance of sexual reproduction and the choice of mate, are discussed.

Light and electron-microscopic observations of the blood-cells (hemocytes) of the wax-moth Galleria mellonella showed that hemolymph coagulation was initiated by the rapid release of material from the granular cells. During incubation for short terms in vitro these cells showed progressive degranulation as material derived from the granules was discharged into the hemolymph. Attempts to determine the nature of this material by staining with ruthenium red proved mainly unsuccessful. When challenged with bacteria in vitro the granular cells failed to phagocytose these particles and instead the bacteria became embedded in the granular material surrounding these cells. The mode of coagulation reported here is compared with previous reports of the role of invertebrate hemocytes in hemolymph clotting.

In Arenicola marina (L.), in vitro attachment and phagocytosis of bacteria appears to be serum-independent and thus, non-specific. Mixtures of Gram+ve and Gram-ve bacterial species (Gram+ve/-ve, of two Gram+ve (Gram+ve/+ve) and of two Gram-ve bacterial species (Gram-ve/-ve)were overlaid on coelomocyte monolayers, in order to introduce competition for cell surface 'receptors'. An enhanced total phagocytosis was only recorded in the Gram+ve/-ve mixtures in comparison with their controls. The results are interpreted in terms of coelomocyte subpopulations and indicate that A. marina coelomocytes may possess specific carbohydrate/glycoprotein determinants capable of recognizing basic carbohydrate differences between the cell walls of Gram+ve and Gram-ve bacteria.

In preliminary experiments, integumental grafting was conducted on a range of insect species in order to elucidate levels of insect immunorecognition. Subsequently, however, only Blaberus craniifer and Extatosoma tiaratum were utilised for detailed study because of their high survival rates and longevity. In both host species, most autografts and allografts rapidly healed and were accepted while xenografts usually became melanized and were rejected. A significant correlation was obtained between the mean rejection times, which ranged from 8.0 +/- 1.8 days to 28.2 +/- 6.6 days, and the phylogenetic relatedness of the xenograft donors to the host species. A preliminary histological investigation revealed the accumulation of blood cells below the grafts, a process that was much more marked with xenografts than with autografts.

The plasmatocytes are the major phagocytic blood-cell type in the haemolymph of the wax-moth, Galleria mellonella. In the present study, these cells were allowed to attach to tissue culture dishes for 1 h, rinsed and then incubated with latex beads for up to 72 h. These cells were then fixed for routine transmission electron microscopy and acid phosphatase cytochemistry. Intracellular latex particles were found in tight, ill-defined phagosomes, which were often clearly associated with the Golgi complexes of the plasmatocytes. Fusion of both primary lysosomes and multivesicular bodies with the phagosomes occasionally occurred and this resulted in the accumulation of an acid phosphatase positive reaction product around the test particles. Subsequent experiments showed that this acid phosphatase activity was mainly associated with the primary lysosomes. The results of the lysosome/latex interactions are compared with those obtained from similar studies on the digestive mechanisms in other phagocytes.

Cellular aggregates or nodules which formed in Galleria mellonella larvae in response to injections of killed Bacillus cereus were examined during their first 24 hr of development. The initial aggregation stage was very rapid, occurring within 1 min, and was brought about by the release of an adhesive flocculent material by granular haemocytes in contact with the bacteria. This material, which subsequently became increasingly electron dense owing to melanin deposition, consolidated and by 5 min ramified all the intercellular spaces. The granular cells degenerated, releasing portions of cytoplasm form their peripheries, and eventurally appeared in the matrix as voids containing loosely flocculent material. No bacterial breakdown was seen, however. From 2-4 hr, plasmatocytes began to encapsulate the necrotic mass, and by 24 hr a complete sheath, composed of three discrete regions of cells, was formed. These findings are discussed in relation to the various proposed mechanisms of nodule and capsule formation.

Examination of whole body homogenates of the hemichordate, Saccoglossus ruber, has revealed the presence of a hemagglutinin with specificity for carbohydrates containing a D-galactose configuration. An analysis of the chemical and physical properties of the agglutinin showed it to be a non-dialysable, heat-labile protein, susceptible to the action of pronase E, but resistant to trypsin. Oxidation by periodate had no effect on agglutination and activity was not dependent upon the divalent cations, Ca2+ or Mg2+. The agglutinin was associated with the proboscis, collar and trunk regions, and was also detected in mucus covering the body surface. A possible role for the agglutinin in the defence system of the animal is discussed.

The mechanism of nonself recognition by the immune system of insects is unknown. In this report the activation of the prophenoloxidase system in the wax moth Galleria mellonella by a microbial product is shown to enhance the recognition of nonself material. These results explain previous observations of the interaction of two different blood cell populations in the cellular defense reactions of insects.

The effect of naturally occurring haemagglutinins on the in vitro phagocytosis of sheep erythrocytes by the blood cells (haemocytes) of Clitumnus extradentatus and Periplaneta americana was studied. The results showed that the haemagglutinins in both species failed to act as opsonins. Indeed, in some instances, incubation of erythrocytes in haemolymph resulted in less avid ingestion as compared with the saline-incubated controls. This reduced phagocytosis was probably caused by the clumping of erythrocytes on the haemocyte monolayers, leaving fewer single red cells available for uptake. The possible roles of these erythrocyte agglutinins in the host defence systems of insects are discussed.

The aim of this work is to study, characterize, and compare different morphological types of hemocytes of Glossina austeni, G. morsitans, Calliphora erythrocephala, Stomoxys calcitrans, Lucilia sericata, Aedes aegypti, and Culex quinquefasciatus. This information is intended to provide a basis for future studies of the cellular defense mechanisms of these dipterans. Seven morphological types of hemocytes were identified by phase-contrast optics: prohemocytes, plasmatocytes, thrombocytoids, granulocytes, adipohemocytes, oenocytoids, and spindle cells of various sizes. Adipohemocytes are difficult to distinguish from both fat body cells and granulocytes. All seven cell types are not present in every species. For example, thrombocytoids and spindle cells were not found in A. aegypti or C. quinquefasciatus, and oenocytoids were observed only in A. aegypti, C. quinquefasciatus, and in C. erythrocephala. In addition to the hemocytes, fat body cells and nephrocytes are also freely present in the hemolymph of some species but may have gained access to the blood during the bleeding process. With electron microscopy and with thick plastic sections of G. austeni hemolymph, only nephrocytes, plasmatocytes, granulocytes, and spindle cells were identified with any certainty, and the large spindle cells are morphologically very different from those found in the other dipterans. They are rigid cells supported by microtubules running throughout their entire length. These hemocytes are present in large numbers only in newly emerged flies, they are absent in larvae and young pupae, and are rare in old adults. Their disappearance from the hemolymph of newly emerged Glossina appears to be a result of phagocytosis by the plasmatocytes.

The SOS DNA repair system is induced in bacteria treated with 4-quinolones. However, whether the response exacerbates or repairs the damage caused by these drugs is still unclear. The recA13 and the recB21 mutations impair recombination repair and render bacteria unable to induce the SOS response when treated with nalidixic acid or other agents that affect DNA synthesis. However, UV treatment induces the SOS response in recB21 mutants but not in recA13 mutants. Both these mutants are hypersensitive to nalidixic acid and, therefore, either recombination repair or SOS repair would appear to repair DNA damage caused by the drug. However, since the lexA3 mutation (which also renders bacteria incapable of inducing the SOS response without affecting recombination repair) had no effect on the susceptibility of bacteria to nalidixic acid, the SOS response neither contributes to nor repairs DNA damage caused by the drug. Consequently, it would seem that the hypersensitivity of the recA13 and recB21 mutants to nalidixic acid is due to their deficiency in recombination repair. This view was confirmed by testing a recA430 mutant that is recombination-repair proficient but SOS repair-deficient and finding it to be no more sensitive to nalidixic acid than its parent. Thus it would appear that, although induced by nalidixic acid treatment, the SOS DNA repair system does not play any role in bacterial responses to the damage caused by the drug. In contrast, the recombination repair system does repair damage caused by nalidixic acid.

Although the origin and evolution of vertebrate blood cells is conjectural and controversial, it is, nevertheless, of great interest, particularly to comparative immunologists seeking an underlying trend in the phylogenetic development of immunity. This review examines possible relationships between invertebrate and vertebrate blood cells and describes some of the original concepts put forward to account for blood cell evolution.

The agglutinating and antibacterial activities of mucus from marine invertebrates are described. Mucus from eleven species was tested for the presence of haemagglutinins using a range of vertebrate erythrocytes. Mucus from Metridium senile, Ophiocomina nigra and Branchiostoma lanceolatum yielded positive results and was investigated further. The haemagglutinin of M. senile was purified, characterized and shown to be mannose-specific. Mucus from this animal was subsequently assayed for antibacterial activity using seawater isolates and verified marine and terrestrial cultures. Only two test organisms were inhibited, while the majority showed significantly enhanced growth. Possible functions of the mucus barrier in marine invertebrates are discussed.

The activity of ten different 4-quinolone antibacterial agents against Escherichia coli KL16 was investigated at four pH values between 5.6 and 8.3, which is the pH range of urine. It was found that pH affected the minimum inhibitory concentrations of all the drugs, but the nature of the substituent at the C7 position of each 4-quinolone governed how its activity was altered by the hydrogen ion concentration. When a 4-quinolone possessed a piperazine group at its C7 position its activity became progressively less as the pH fell. However, all drugs that lacked a C7 piperazine, irrespective of the nature of their substituent at this position, exhibited a progressive increase in activity as the pH was reduced. In addition it was found that, excepting cinoxacin, a urinary concentration of magnesium generally caused a further antagonism of the activity of the nine other 4-quinolone antibacterials at all pH values tested.

Larvae of the cabbage white butterfly, Pieris brassicae, were reared on a semisynthetic diet with or without 20 ppm of the herbicide 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) and using three assays the sera were subsequently tested for natural antibacterial activity against Bacillus cereus, Escherichia coli K12, and Micrococcus luteus. These assays showed that exposure of larvae to 2,4,5-T lowered the antibacterial activity of the serum against E. coli and M. luteus compared with control animals. Spectrophotometric tests for the presence of a lysozyme-like principle in the serum also revealed similar trends with a significant loss of enzyme activity in 2,4,5-T-treated insects. Overall total serum protein levels of control and 2,4,5-T-treated insects were similar, suggesting a specific effect of the herbicide on certain serum components such as lysozyme. The possible mode of action of the herbicide on production of antibacterial factors is discussed.