- Volume 13, Issue 4, 1980

PLATES XXX-XXXII

THE ENZYME urease is thought to play a major role as a virulence factor in urinary-tract infections with Proteus (Braude and Siemienski, 1960) although it may not be the only factor (Eudy, Burrous and Sigler, 1971). Urease degrades urea with release of ammonia which may cause damage and death to the renal epithelium, inactivation of complement, and conditions favouring the development of renal stones. Many of these effects can be prevented by the administration of urease inhibitors (Griffith and Musher, 1973; Griffith, Musher and Campbell, 1973; Aronson, Medalia and Griffel, 1974; Musher et al., 1975).

Phillips (1955) found wide variation in the pathogenicity of different strains of Proteus for the mouse kidney in vivo, and Senior (1979) has shown that particular proticine-production/sensitivity (P/S) types of P. mirabilis have a special affinity for the urinary tract whereas other P/S types and other Proteus species are seldom incriminated in serious urinary-tract infections. The aim of this study was to determine whether or not these differences in pathogenicity among strains of Proteus were associated with differences in the types of ureases produced by these strains.

Sodium dodecyl sulphate extracts of the reference strains Mus 127, Castellón 3 and Arborea of the Ballum agglutinogenic serogroup of Leptospira interrogans (the species of pathogenic leptospira), of strain Patoc 1 of the saprophytic species of L. biflexa, and of strain 3055 of Mini serotype, the sole representative of L. Mini, were each fractionated by ultracentrifugation in a sucrose density gradient into 10 fractions. The fractions were tested by complement fixation and immunodiffusion against the sera of animals during the process of immunisation and during the course of naturally occurring infections.

The fractions could be divided into three main pools of serological activity: pool I (fractions 1,2 and 3), pool II (fractions 4, 5 and 6), pool III (fractions 7,8 and 9). Pool I was species/genus specific; fraction 1 tended to be species specific while fraction 2 reacted with antisera to all strains whether pathogenic or saprophytic. Pool II was serogroup specific and reacted only with antisera to members of the same serogroup. Pool III was serotype (serovar) specific and revealed the identity of the infecting strain at an early stage of infection.

Local respiratory-tract infection was produced experimentally in guinea-pigs by intranasal instillation of a suspension of parainfluenza virus type 3. Histologically, interstitial pneumonitis developed within 10 days and persisted for at least 70 days.

Cell-mediated immunity was measured at intervals for 70 days after infection. Dermal reactivity could not be elicited. Leucocyte-migration inhibition and macrophage-migration inhibition were increased. Macrophage aggregation was present. Increased cell-mediated immunity could be transferred from infected donor animals to normal recipient animals by adoptive spleen-cell transfer even 60 and 70 days after infection.

Of 30 prednisolone-treated mice challenged intravenously with Mycobacterium haemophilum, 12 developed skin lesions, primarily on the ears, from which the mycobacterium was isolated. These lesions were not observed in mice challenged with M. haemophilum but not given prednisolone.

M. haemophilum infection prolonged the life and reduced the frequency of intercurrent infection with Corynebacterium murium in prednisolone-treated mice.

The ability to form gas in lactose bile-salt broth at 44.5°C (the “faecal coliform” or FC test), growth in nutrient broth at 10°C, indole production and pectin liquefaction were studied in 480 strains of Klebsiella isolated from human and animal infections, from various sites in the hospital environment and hospital food, and from river water and flowers. A positive FC response was correlated inversely with the ability to grow at 10°C. Most strains of human and animal clinical origin were FC positive, whereas strains from water and flowers were mainly FC negative. The frequency of a positive FC response in strains from the hospital environment fell between these two extremes. The production of indole and liquefaction of pectin by klebsiellas was correlated directly with the ability to grow at 10°C and a negative FC response. Nearly all of the strains could be allocated to one of four groups on the basis of these tests.

The capsular serotype, bacteriocine-inhibition patterns and antibiotic sensitivities of the strains were examined. No correlation was evident between the first two properties and klebsiellas from any particular source. Strains of clinical origin were more often resistant to five or more antibiotics than were strains from the hospital environment, which in turn showed a greater frequency of antibiotic resistance than did strains from river water and flowers.

An enzyme immunoassay (EIA) for the capsular polysaccharide of Streptococcus pneumoniae type III was developed and applied to the measurement of this antigen in cerebrospinal fluid (CSF) in an experimental model of pneumococcal meningitis. EIA was performed by a single-antibody sandwich technique in which the globulin fraction of pneumococcal type-specific antiserum (antiserum-globulin) was used to coat the solid phase before antigen attachment and to conjugate with the labelling enzyme, horseradish peroxidase. Under optimum assay conditions EIA detected purified pneumococcal type-III capsular polysaccharide in concentrations as low as 0.15 ng/ml in aqueous buffer. Assayed by EIA, the mean concentration of type-III capsular polysaccharide in CSF of rabbits with pneumococcal meningitis increased exponentially from 24 h to 96 h of infection (range 13.9 ng/ml–62 500 ng/ml). Effective antimicrobial therapy of rabbits with meningitis was associated with a rapid decrease in the CSF concentration of the capsular antigen, although it was still detected in concentration in the range 1–10 ng/ml in 100% of animals treated for 4 days. Thus EIA provides a quantitative and extremely sensitive method of measuring type-III pneumococcal capsular polysaccharide in CSF.

PLATES XXXVII-XXXVIII

THE OCCURRENCE of excess infant deaths during influenza epidemics (Dauer and Serfling, 1961; Wynne Griffith et al., 1972) suggests that influenza plays a greater role in infant death and severe illness than is generally recognised (Nelson et al., 1975; Paisley et al., 1978). Although children are highly susceptible to influenza infection, their symptoms may be milder than in the adult (Douglas, 1975). However, convulsions, croup and pneumonia also occur (Brocklebank et al., 1972; Naude et al., 1974; Spence, Brodie and Masson, 1975; Laraya-Cuasay et al., 1977; Paisley et al., 1978). The pathology of influenza in infants has rarely been reported, and then usually in cases complicated by other disease processes (Louria et al., 1959); by contrast, the findings in adult man (Hers and Mulder, 1961) and in the adult ferret (Francis and Stuart-Harris, 1938) have been well described.

Influenza in the healthy adult ferret is a transient, non-fatal illness, similar to that occurring in the vast majority of human adults (Toms et al., 1976). This similarity of clinical and pathological responses in man and ferret, and the recognition of fatal infection in the fetal ferret (Collie et al., 1978) stimulated the present investigation into the susceptibility of the newborn ferret to influenza.

Some clinically significant anaerobic bacteria were incubated in pure culture in anaerobic jars containing a range of atmospheric concentrations of CO2. Growth of Clostridium perfringens, C. sporogenes and C. septicum was independent of the amount of CO2. Small supplements of CO2 (0.25%) allowed good growth of the majority of anaerobes studied—an observation contrary to established teaching. The exceptions were Fusobacterium necrophorum which showed an absolute requirement for CO2 of at least 1% and B. melaninogenicus which needed an atmospheric content of 10–40% CO2 for optimal growth. The inclusion of CO2 in the anaerobic jar at a final concentration of 10% is to be recommended for all routine isolation procedures.

Tween 80 stimulated growth of Corynebacterium ulcerans but suppressed toxin production. It had no direct effect on preformed toxin. Toxin production was not inhibited by oleic, linoleic, palmitic or stearic acids.

PLATE XL

CORYNEBACTERIUM ULCERANS, which is usually associated with mastitis in cattle, has been reported as a cause of sore throat and tonsillitis in man (Jebb, 1948; Saxholm, 1951; Tomlinson, 1966). Subcutaneous injection of this organism into guinea-pigs produces features, such as haemorrhage and oedema at the site of injection and haemorrhagic adrenals, which are characteristic of diphtheria toxin. Diphtheria antitoxin neutralises the lethal effect of toxin from some strains of C. ulcerans (Fox and Frost, 1974) but not from others (Souckova and Soucek, 1974). In this study the purification and preliminary characterisation of a toxin from C. ulcerans strain 378 is reported.

The survival of gonococci on various materials contaminated with gonococcal pus and stored at room temperature was studied. Gonococci were recovered for up to 3 days from a wide variety of hard and soft materials. It is possible that gonorrhoea is transmitted nonvenereally more often than is usually acknowledged, and these results may have medicolegal and social significance.

The ANM medium of Hafiz and McEntegart (1976) was found to be deficient in ability to support the growth of various strains of N. gonorrhoeae. Strains that did grow required a large starting inoculum which invariably suffered a substantial drop in the number of viable organisms during the first 2 h. Investigation of the various deficiencies led to the development of liquid (G77L) and solid (G77S) media which were shown to have several advantages over existing media for the cultivation of N. gonorrhoeae. These media, which are simple to prepare, do not contain blood or serum and yet gave excellent growth. The inclusion of antibiotics, glucose and phenol red allowed the selective isolation and partial identification of N. gonorrhoeae from routine specimens. These media also gave good results in fermentation tests for the identification of neisseriae.

The relative effect of humidity, temperature and exposure to oxygen on the survival of 11 strains of Bacteroides spp., Fusobacterium necrophorum, Eubacterium lentum, Clostridium ramosum, Peptococcus asaccharolyticus and Peptostreptococcus anaerobius was evaluated. Suspensions of the test strains were applied to membrane filters either in empty sterile petri dishes or on plates of non-nutrient agar to provide dry or moist conditions. The filters were stored aerobically or anaerobically in an atmosphere containing 10% CO2 at either 4°C or 35°C. After holding periods of 0.375−384 h, the filters were transferred to nutrient plates inside an anaerobic glove box. After incubation the growth on the filters was compared with that on control filters that had not been stored. Survival was better at 4°C than at 35°C. For the gram-negative organisms, survival was better under moist than under dry conditions, whereas for gram-positive organisms this finding was sometimes reversed. The results indicate that, generally, anaerobic bacteria kept under aerobic conditions survive best under moist conditions at 4°C.