Counts Of Bacilli Research Articles

Changes in the microbial profile and proximate composition during natural and controlled fermentations of soybeans to produce kinema

The mean viable count of Bacillus subtilis, Enterococcus faecium and Candida parapsilosis increased progressively in fermenting soybeans; the increase was significant at every sampling time interval (8 h) up to 48, 40 and 32 h, respectively. Although acidity increased significantly at almost every 8 h interval, the fermentation was essentially alkaline, causing pH of the beans to rise to about 8·5. With the decline in protein nitrogen content, the non-protein and soluble nitrogen contents increased significantly at almost every 8 h interval. The fermentation process as well as general acceptability of kinema were improved by fermenting sterile beans with B. subtilis DK-WI at 45°C. This led to a more desirable fermentation within a much shorter period, compared with the natural fermentation. The rate of increase of Bacillus count in monoculture fermentation was much faster than that in natural fermentation. The Bacillus count in kinema produced by monoculture fermentation was 6·4 times higher than that of naturally fermented kinema. The content of free fatty acids of the Bacillus-fermented kinema was 140% less than that of the naturally fermented one. However in respect of other compositional parameters, the two types of fermentation were similar.

Bacillus spore contamination associated with commercial bread manufacture

Rope spoilage of bread by Bacillus subtilis and B. licheniformis causes economic losses to the baking industry. The Bacillus spore contamination profile of a typical brown bread manufacturing line in a commercial bakery was assessed by four replicate microbiological surveys to determine numbers, types and sources of Bacillus spores. High Bacillus spore counts were consistently obtained from yeast, crumbs and premix. Food contact surfaces did not contribute noticeably to spore counts. The initial spore count of dough after mixing was 2·6 log cfu g -1 and increased by 0·5 log cfu g -1 before baking. Baking decreased the Bacillus spore count by 1 log cfu g -1. Baked bread stored at 30°C yielded Bacillus counts of c. 10 7 cfu g -1 after 3 days' storage. Three hundred randomly selected Bacillus isolates from raw materials, dough, food contact surface and bread samples were characterized as 41·5% B. licheniformis, 38·4% B. subtilis, 12·0% B. megaterium, 3·9% B. pumilis, 2·5% B. laterosporus and 1·1% B. cereus.

Efficacy of intermittent pyrazinamide in experimental murine tuberculosis

CFLP mice were infected intravenously with Mycobacterium tuberculosis strain H37Rv and the progress of chemotherapy was followed by counts of viable bacilli in the lung and spleen. After spleen counts had reached log10 7.0, 12 experimental groups, each containing 10 mice, were treated for 8 weeks with pyrazinamide (PZA) given in mean daily dosages of 100, 200 or 400 mg/kg/day, with the interval between the doses within each dosage group being 1, 2,4 or 8 days. All mice were also given 25 mg isoniazid/kg daily. An increase in the mean daily dosage from 100 mg PZA/kg to 400 mg PZA/kg resulted in a decrease of spleen viable counts at the end of treatment from log10 4.2 to log10 3.8. The organ counts, averaged over the full dosage range, were little altered by spacing out the interval between doses from 1–4 days, while increasing dose size proportionately: the counts with low mean dosages tended, however, to decrease (indicating improved efficacy) while those with high mean dosages increased (P<0.001). Counts increased when the interval was 8 days. Spacing out the doses while keeping the dose size constant resulted in progressive loss of efficacy. These findings suggest that, if PZA is given intermittently, the size of the dose should be increased, though not quite proportionately, to maintain full efficacy. Even with such an increase in dose, however, once weekly treatment would be less effective.

Relative permissiveness of macrophages from black and white people for virulent tubercle bacilli

Epidemiological, clinical, and histopathological evidence suggests that black people are more susceptible to tuberculosis than are white people. The cellular basis of this putative susceptibility was investigated in vitro by comparing responses of blood-derived macrophages from black and white donors to experimental infection with virulent tubercle bacilli. Phagocytes from pairs of black and white donors were infected. The uptake and replication of the tubercle bacilli in these cells were measured by microscopic counts and by CFU counts of bacilli at 0, 4, and 7 days. The effects of donor serum, of 1,25-(OH)2-vitamin D3, and gamma interferon on the infection also were studied. Black-donor phagocytes killed more bacilli during phagocytosis than white-donor phagocytes did. However, the bacilli grew consistently and significantly faster in successfully infected macrophages from black than from white donors, especially in the presence of black-donor serum. 1,25-(OH)2-vitamin D3 gave significantly less protection against tubercle bacilli to macrophages from black donors than to macrophages from white donors. The permissiveness of the macrophages from the two races was affected equally by gamma interferon. These results demonstrate some inherent and environmental liabilities in the monocytic phagocytes and serum of black people compared with white people, which may contribute to their greater susceptibility to tuberculosis.

Pathogenicity of Mycobacterium avium for human monocytes: absence of macrophage-activating factor activity of gamma interferon

Mycobacterium avium is a frequent opportunistic pathogen in the acquired immunodeficiency syndrome (AIDS). We compared 12 strains of M. avium in an in vitro model of pathogenicity. Peripheral blood-derived monocytes from healthy individuals were infected with M. avium in vitro. Bacterial uptake and intracellular replication were assessed by microscopic count of acid-fast bacilli and CFU of bacteria, respectively, in lysed monocytes. The CFU assay showed that among five AIDS-associated strains, only one replicated in monocytes. Two of seven non-AIDS-associated strains replicated intracellularly. In addition, we examined the effect of gamma interferon (IFN-gamma) on M. avium infection. IFN-gamma treatment of monocytes decreased phagocytosis and had no effect on the intracellular replication of M. avium. Thus, most strains of M. avium do not multiply within monocytes from healthy individuals and IFN-gamma does not have macrophage-activating factor activity for M. avium infection of human monocytes.

Cytokine regulation of the intracellular growth of Mycobacterium paratuberculosis in bovine monocytes.

In this study we examined the influence of various crude and recombinant cytokines on the ingestion and intracellular survival of Mycobacterium paratuberculosis within bovine monocytes and monocyte-derived macrophages. Cytokine pretreatment had little effect on the ingestion of M. paratuberculosis by bovine monocytes and macrophages. Monocytes that were continuously incubated with virus-induced crude bovine interferon (100 U) or recombinant bovine alpha interferon (100 U) significantly restricted the subsequent intracellular growth of M. paratuberculosis, as determined by microscopic counts of acid-fast bacilli and by recovery of CFU from lysed monocyte monolayers. In contrast to their effects on freshly adherent monocytes, these cytokines had little effect on the growth of M. paratuberculosis within monocyte-derived macrophages. In two separate experiments, we also observed inhibition of bacillary growth in monocytes treated with unpurified recombinant human granulocyte-macrophage colony-stimulating factor. Conversely, intracellular growth of M. paratuberculosis was enhanced in monocytes that were pretreated with culture supernatants from M. paratuberculosis-stimulated peripheral blood mononuclear cells obtained from an immunized calf. The growth-enhancing activity of these supernatants was labile at pH 2.0, suggesting a role for gamma interferon; however, subsequent experiments indicated that recombinant gamma interferon alone neither enhanced nor restricted intracellular bacillary growth. To determine the possible contributions of monocyte oxidative activity to cytokine-induced bacteriostasis, we compared the release of superoxide anion from cytokine-treated and control monocytes. No obvious relationship was observed between the release of superoxide anion and the subsequent intracellular fate of the bacilli.

The role of host factors for the chemotherapy of BCG infection in inbred strains of mice.

Host factors have previously been considered to play a role in the efficacy of "incomplete" antituberculous chemotherapy. We investigated this aspect using a model of intravenous infection with M. bovis-BCG in three inbred strains of mice. Viable splenic counts of bacilli (CFU) were monitored following a three week regimen of rifampicin (R) and isoniazid (H) given either immediately or from three weeks after infection. At the end of the immediate (0-3 weeks) therapy the spleens of all mice were sterile and only a minority of animals had demonstrable CFU's at 23 weeks after infection. However, following the delayed-onset (3-6 weeks) therapy we found a pronounced relapse of BCG growth which was about 10 times higher in CBA/Ca and BALB/C than in C57B1/6 mice. These results indicated that host immunity which developed during the first three weeks after infection may have aggravated the relapse of bacterial growth following chemotherapy. This interpretation was corroborated by the finding that the immunosuppressive drug cyclosporin given concurrently with R/H reduced the splenic viable counts of BCG.

Effectiveness of ofloxacin against Mycobacterium tuberculosis and Mycobacterium avium, and rifampin against M. tuberculosis in cultured human macrophages.

Ofloxacin (OFL) is a new broad-spectrum drug with potentially valuable antimycobacterial activity. It was tested for ability to inhibit virulent tubercle bacilli (TB) and virulent Mycobacterium avium in cultured human macrophages (MP). The first-line antituberculosis drug rifampin (RMP) also was tested against TB in MP. The drugs were added to the MP cultures immediately after infection or 2 days later. Antimicrobial inhibition was measured by colony-forming-unit (cfu) counts of bacilli from lysed samples of the infected MP taken at zero, 4, and 7 days after infection. Drug inhibition of the bacteria in vitro in 7H9 broth also was measured. OFL showed the same minimal inhibitory concentration (MIC) of 1.25 micrograms/ml against TB in vitro and in MP. At 2 micrograms/ml or more it killed TB in vitro and in MP. It was equally effective in MP against initially nonmultiplying TB and MP-established TB, which were multiplying exponentially in the MP. OFL had the same MIC in vitro against M. avium but was ineffective against both M. avium in MP and M. avium isolated directly from MP at as much as 8 micrograms/ml. The MIC for RMP against TB in MP was 0.1 microgram/ml, and TB in vitro 0.02 microgram/ml. At 0.5 microgram/ml or greater, it killed TB in MP. RMP killed TB in MP rapidly, whereas OFL killed them slowly. These results confirm initial clinical evidence, as published by others, that OFL should be a useful antituberculosis drug. However, they suggest that it may not be very effective against MA infections.(ABSTRACT TRUNCATED AT 250 WORDS)

Selection and counting of aerobic gram-negative bacilli in saliva by the spiral system.

A four-month pilot study involving 100 volunteers, 100 hospitalised patients not on antibiotics and 100 patients on antibiotics was performed using a non labor-intensive system involving inoculator, laser colony counter and 14-cm petri dishes containing MacConkey medium to determine the number of aerobic gram-negative bacilli present in saliva. All colonies greater than or equal to 0.75 mm in diameter were found to be aerobic gram-negative bacteria. This was also valid for Pseudomonas aeruginosa, but only after incubation for 48 h. This study showed that the novel and practical technique used can be applied to a large number of saliva specimens, and antibiotics have limited impact on the buccal microflora (intensive care unit excluded).

Inhibition by Pyrazinamide of Tubercle Bacilli within Cultured Human Macrophages1–3

Pyrazinamide (PZA) is a unique antituberculosis drug because it is effective in vivo but not in mediums commonly used to culture tubercle bacilli. Consequently, it was employed to test the validity of an in vitro macrophage model of human tuberculosis for value as a correlate of clinical events. The drug was as active in the macrophage model as it was clinically, inhibiting virulent tubercle bacilli at concentrations at 20 micrograms/ml or higher. By contrast, it was ineffective in 7H9 bacteriologic culture medium, even at concentrations as high as 2,560 micrograms/ml. It could be either bacteriostatic or bactericidal against intramacrophage tubercle bacilli, depending on its concentration, the donor of the macrophages, and the length of exposure of the infected macrophages to the drug. The data presented suggest that the clinical effectiveness of PZA is determined by a complicated self-modulating sequence of interactions between it, tubercle bacilli, and host macrophages. Specific evidence was found that tubercle bacilli may replicate within human macrophages in non-acid-fast form, thus indicating that colony-forming unit counts are inherently more accurate than acid-fast bacilli counts in these experiments, and also suggesting that important changes in bacillary cell wall composition may occur among tubercle bacilli within infected human macrophages.

Changes in Microbial Population and Growth of Bacillus cereus During Storage of Reconstituted Dry Milk

Eight brands of retail nonfat and whole dry milk were reconstituted and analyzed for changes in the predominant bacterial population and for the proliferation of Bacillus cereus throughout storage at 30, 20, and 5°C. All brands yielded similar results. Bacillus and Micrococcus predominated in the freshly reconstituted milk. During storage at 30°C, the Bacillus population proliferated initially, but was gradually replaced by enterococci. At the time of spoilage, Bacillus counts had dropped by several orders of magnitude. The proportion of Micrococcus gradually declined. B. cereus counts reached hazardous levels as early as 10 h after reconstitution, which was before spoilage was evident. Similar changes occurred in reconstituted milk stored at 20°C, but the time course for the changes was longer, and the Bacillus counts did not decline as rapidly as they did at 30°C. Again, counts of B. cereus reached hazardous levels before the milk showed signs of spoilage. At 5°C, the milk showed no signs of spoilage for 4 to 5 weeks. Bacillus constituted more then 90% of the bacteria isolated after the first week. Bacillus counts continued to increase slowly, but the relative proportion decreased as the gram-negative rods, especially Enterobacter, proliferated. B. cereus never reached numbers great enough to cause disease. The results revealed that the microbial profile of reconstituted dry milk changed significantly over time and that the temperature of storage determined the eventual microbial composition. The results also showed that B. cereus is an omnipresent health hazard in reconstituted milk that is not properly refrigerated.

Microbiological, physical and chemical studies of Livestock-farm water in Taiwan

The properties of drinking water from 42 hog farms, including pH, color, turbidity, total soluble solids, concentrations of ammonium nitrogen, nitrate nitrogen, nitrite nitrogen, chlorine, fluorine, sulphate, calcium, copper, zinc, iron, manganese and chromium were investigated. The total number of coliforms, fecal coliforms, fecal streptococci and standard plate counts of both drinking water and waste water were also studied. It was found that all the waste water and some of the drinking water was polluted. The major pollutant was hog feces. The culture counts of aerobic Gram-negative bacilli of fresh feces and fecal polluted water on one hog farm were examined in detail by using selective media. There were 364 strains of pure cultures isolated from MacConkey agar. The biochemical characteristics of the isolates on the triple sugar iron agar, Simmon's citrate agar, Christensen's urea agar, sulfide-indole-motility medium, Voges-Proskauer broth, Moller's ornithine decarboxylase medium, etc., were investigated. Microbial identification was carried out according to a computer-coding system based on the characteristics shown in the above media. The identified species were Escherichia coli (71%), Enterobacter agglomerans (6%), Enterobacter cloacae, Citrobacter freundii, Citrobacter diversus, Citrobacter amalonaticus, Aeromonas hydrophila, Arizona hinshawii, Vibrio sp., Hafnia alvei (Enterobacter hafniae), Chromobacterium violaceum, Yersinia enterocolitica, Plesiomonas shigelloides, Serratia liquefaciens, Pectobacterium sp., Proteus mirabilis, Edwardsiella tarda, Vibrio vulnificus, Provindencia rettgeri, Provindencia alcalifaciens, Proteus vulgaris, Klebsiella ozaenae, Klebsiella sp., Shigella sp., Morganella morganii, Serratia marcescens and Vibrio (EF-6) group F. The significance of these isolates for public health is discussed.

The effect of "humanised" milks and supplemented breast feeding on the faecal flora of infants.

SUMMARY Faecal specimens from 47 babies were examined each week during the first 6 weeks of life. Thirteen infants received breast milk alone, nine received Cow and Gate Premium milk, 10 received Scientific Milk Adaptation Gold Cap milk, and 15 received breast milk and in the first week of life a supplement of modified cows' milk. An acetate buffer was demonstrated in the faeces of >60% of breast-fed infants during the first 4 weeks of life. No buffer was demonstrated in the faeces of either bottle-fed group over the same period. Only 20% of breast-fed infants receiving a supplement produced an acetate buffer in the 1st week, but this figure increased to 60% by the 4th week. Breast-fed babies produced faeces with a low pH (5·1-5·4), high counts of bifidobacteria and low counts of coliforms, bacteroides and clostridia; with the exception of acetic acid there was a notable absence of most volatile fatty acids from these faeces. Infants fed Premium milk produced faeces with a high pH (5·9-8·0) and a relatively high bifidobacterial count in the 1st week; thereafter no bacterial species predominanted. A wide variety of volatile fatty acids was present in these infants' faeces. Infants fed Gold Cap milk produced faeces with a high pH (6·4-8·2) and a wide variety of volatile fatty acids; the counts of coliform bacilli and putrefactive bacteria were high, whilst counts of bifidobacteria were low. Breast-fed infants who received a supplement in the 1st week produced acid faeces, but the pH was higher than that of infants fed breast milk alone; the bacterial flora was putrefactive for the first 2 weeks but gradually became fermentative, and whilst several volatile fatty acids were detected in the faeces in the 1st week they gradually decreased during the following 5 weeks.

Systemic Mycobacterium lepraemurium infection in mice: differences in doubling time in liver, spleen, and bone marrow, and a method for measuring the proportion of viable organisms in an inoculum.

Counts of acid-fast bacilli were made on homogenates of whole liver, whole spleen, and two femurs of CBA mice killed at various time intervals after intravenous infection with Mycobacterium lepraemurium. The growth curves so obtained showed that the bacillus multiplied faster in bone marrow than in liver or spleen. No evidence of redistribution during the early part of infection was obtained. The time of appearance of significant numbers of bacilli (10(7)) in the bone marrow was used to make estimates of viability of M. lepraemurium suspensions. Several applications of the techniques described are discussed.

Drug-resistant pulmonary tuberculosis treated with ethambutol, a rifamycin and a riminophenazine (B663)

TUBERCULOSIS associated with tubercle bacilli tha t have become resistant to the standard and reserve drugs may be arrested by a combination of ethambutol, a rifamycin, and the riminophenazine B663. In 1948 a man of 18 developed tuberculosis in the right lung and later in the left lung. A pneumothorax, lobectomy, thoracop!asty, and a variety of chemotherapeutic regimes checked the progress of the disease only intermittently. By 1965 the remains of the left lung and the apex of the right lung were cavitated, and the tubercle bacilli were resistant to streptomycin, PAS, isoniazid, cycloserine, ethionamide, pyrazinamide, viomycin, kanamycin, capreomycin, thiosemicarbazone, and isoxyl. They were susceptible to ethambutol (resistance ratio 4/4), rifamide (o.25/o.25) , and the riminophenazine B663 (o.I2/O.I2). The patient was given ethambutol, 1.6 g daily for I I weeks and thereafter 1 g daily; rifamide i g intravenously on 5 days weekly for 8 months and then twice weekly for 6 months, followed by the oral rifampicin 45 ° mg on 4 days weekly for 4 months; and the riminophenazine B663 3oo mg daily. The most striking effect was the rapid disappearance of tubercle bacilli from the sputum. Before starting treatment, there were i to io tubercle bacilli per I ]7th oil immersion field in smears of untreated sputum; and 4 to 5 millions per ml of sputum as estimated by culture (viable count). Sputum was examined weekly after chemotherapy had been started. The viable count of tubercle bacilli was 2oo at seven weeks, and no growth was obtained at eight weeks; and tubercle bacilli had disappeared from the smears at i o weeks. At this time the sputum ceased to be purulent and became mucoid. Large numbers of tubercle bacilli reappeared in the smear at 44 weeks but they did not grow on culture. A single colony of tubercle bacilli was obtained from the sputum at 46 weeks. It was susceptible to ethambutol, rifamide, and B663. Salmon-pink streaks were seen in the sputum for a time 12 months after treatment had begun. Wet films of these streaks showed the presence of masses of red, spicular crystals of B663. Eighteen months after the start of treatment the patient developed an acute respiratory infection associated with a raised serum titre to adenovirus and died. Post mortem 23 samples were taken from lung tissue, cavity walls and contents, calcified masses, and hilar glands. The deposits from the homogenates

Multiplication of Mycobacterium leprae in the mouse foot-pad. 1. Thermoresistance of Mycobacterium leprae

By the aid of the mouse foot-pad passage technic as a means of evaluation for the multiplication of Mycobacterium leprae, observation was made on the survival of this organism at different temperatures. Strain B2409, isolated by Dr. Shepard and maintained at the author's laboratory, and strain Yamamura, isolated at the same laboratory, were employed as sources of M. leprae. The procedures used for inoculation, harvesting, and quantitative evaluation were essentially the same as those described by Shepard.Hanks' balanced salt solution (BSS) was used for preparing a bacillary suspension and its dilutions. The mice used were of the CF # 1 strain.Storage of bacillary suspension at 5°C: A suspension containing 1.6×105 acid-fast bacilli (strain B 2409) per ml in BSS was stored in a refrigerator at 5°C over a period from 24 hours to 2 weeks. At certain intervals of time during storage, 0.03ml of bacillary suspension, containing 4.8×103 organisms, was inoculated into each mouse at the left hind foot-pad. Harvesting was made on the 120th, 180th, 300th. and 365th day after inoculation. The number of acid-fast bacilli in the foot-pad was counted by the microspot method.There was no significant change in viability of the organisms among the samples at 24, 48, 72, and 96 hours of storage at 5°C. A considerable reduction in their viability, however, was observed in the sample at 7 days of storage.Storage of bacillary suspension at -25°C: A suspension of M. leprae (strain Yamamura) containing 4.8×103 bacilli per 0.03ml was stored in a deep freezer at -25°C over a period from 24 hours to 12 weeks. Then it was inoculated into mice at the foot-pad. The counting of bacilli was performed in the same way as mentioned above. The viability of M. leprae was not significantly affected by freezing and storage at -25°C within 12 weeks.Inactivation of the organism at 60°C: A suspension of M. leprae (strain B2409) containing 4.8×103per 0.03ml was kept in a water bath at 60°C for 5 to 60 minutes. Then 0.03ml of this suspension was inoculated into mice at the foot-pad. As a result, the viability of M. leprae was reduced gradually by the prolongation of time for treatment at 60°C, and completely lost by heating for 30 minutes.

An Assessment of Microbiological Quality of Some Commercially Packed and Fresh Fruit Juice Available in Dhaka City: A Comparative Study

Fruit juices are regarded as the most preferred non-alcoholic beverage worldwide to all age groups. In general, microbial growth in fruit juice is restricted by using preservatives and also through other environmental factors during production. Still, the presence of some indicator organisms in fruit juice is responsible for causing food-borne diseases and even death. So, from public health point of view, it is quite important to know the microbiological quality of the fruit juices available in the market. Primary objective of this study was to assess the microbial quality of fresh and commercially packed available juices collected from different locations of Dhaka city. A total of six fresh juice and nine commercially packed juice samples were collected. Standard culture techniques were followed to assess total viable count (TVC), total staphylococcal count (TSC), total Bacillus count (TBC) and total fungal count (TFC) on different culture media. The TVC varied from the range of 102 to 105 cfu/ml with the highest of 2.4 x 105 cfu/ml. A large number of staphylococci and Bacillus was also found from several samples. Total coliform and fecal coliform was found in six and five (out of fifteen) samples, respectively. Among total coliforms, Klebsiella, Enterobacter along with E. coli were detected. From all the assessment it was determined that the microbial quality of commercially packed juice was fairer than that of fresh juice collected from local market. DOI: http://dx.doi.org/10.3329/sjm.v1i1.9097 Stamford Journal of Microbiology, Vol.1(1), July 2011, p.13-18

マウス足底部におけるM.lepraeの増殖

The transmission of M. leprae into experimental animals had been studyed by means of the inoculation into foot-pads of mice since 1964, and it was indicated that M, leprae could multiply successfully in the tissue of mice foot-pads, as reported by C.C. Shepard. In our laboratory, M. leprae was isolated from leproma of Japanese leprosy patients of lepromatous type by using the mice of some Japanese strains. At present, 9 strains of M. leprae are maintained in more than fourth generation and 7 strains of M. leprae are kept under investigation. In 9 strains of M. leprae, 7 strains are isolated from untreated patients and 2 strains are isolated from treated patients. Very easy method was used for the counting of acid-fast bacilli; The emulsion of tissue of mouse foot-pad was made by using the pestle arid mortar, and one drop of this emulsion was located on each slide-glass by using the pestle. The number of acid- fast bacilli in this smear was counted microscopically and the average bacillary count could be obtained.In this studies, it was found that the multiplication of bacilli was indicated by the appearance of globi-like findings gathered each other with bacilli and that this growth was limited in the foot-pad of mouse. The transmission of M. leprae into mice of next generation may be tried at about 6 months after the infection and the bacillary number of more than 104 in each foot-pad of mouse may be required in subtransmission, because many killed bacilli might be contained in this emulsion of tissue of mouse foot-pad.The multiplication of M. leprae was increased by the administration of Cortisone, but this could not be influenced by the injection with Mitomycin C, 6-Mercaptopurine and 2-Mercaptoethanol. However, the subcutaneous injection of Adjuvant into the foot-pads of mice inhibited the multiplication of M. leprae.

Host-parasite relationships in experimental airborne tuberculosis. I. Preliminary studies in BCG-vaccinated and nonvaccinated animals.

Smith, D. W. (University of Wisconsin, Madison), E. Wiegeshaus, R. Navalkar, and A. A. Grover. Host-parasite relationships in experimental airborne tuberculosis. I. Preliminary studies in BCG-vaccinated and nonvaccinated animals. J. Bacteriol. 91:718-724. 1966.-Previous studies from this laboratory on immunogenicity and allergenicity of defatted mycobacterial vaccines involved subcutaneous challenge of guinea pigs and killing of the animals 6 weeks later to evaluate the amount of disease. This type of experiment has discontinued in this laboratory in favor of an airborne challenge type of experiment, with the advantages that animals can be challenged with small numbers of bacilli by a natural route, and the number of primary lesions, the rate of spread from those lesions, and the rate of bacillary multiplication can be used to evaluate protection. Experiments to determine uniformity of infection showed that a fair degree of uniformity resulted when seven guinea pigs were exposed simultaneously, and were studied 3 weeks later to determine numbers of primary lesions and bacilli in the tissues. A less satisfactory degree of uniformity was obtained when more animals were exposed at one time. BCG-vaccinated and nonvaccinated animals were studied to determine the earliest time and the optimal time for killing the animals to detect the effects of vaccination. In guinea pigs, the degree of protection assessed by lesion counts is time-dependent, but the degree of protection assessed by viable counts of bacilli in the tissues was relatively constant 3 to 12 weeks after infection. Mice vaccinated subcutaneously with BCG were not protected against infection at any interval between 2 and 19 weeks. Guinea pigs vaccinated subcutaneously with the same lot of vaccine were protected as judged by counts of viable bacilli in the tissues 3 weeks after infection.

The lethal effect of cotton-wool lipid on tubercle bacilli in acid conditions and its prevention by surface-active agents

Long-chain fatty acids are not appreciably bactericidal to tubercle bacilli at neutral pH, but become so even in slightly acid solution. In this way tubercle bacilli, normally resistant to 0·1 N hydrochloric acid (pH 1·0), are rapidly killed at this pH if traces of fatty acid are present, such as can be found inside test tubes that have been plugged with cotton wool and sterilized by dry heat.The lethal effect is largely prevented by non-ionic, cationic, and to a lesser extent anionic detergents, which probably take up the fatty acid into the micelles so that it can no longer attack the bacilli.The various fatty acids identified in the deposit from heated cotton wool were tested individually and found to be bactericidal. They comprised the usual C12to C18saturated acids and also oleic acid. The latter, however, showed no particular lethal toxicity in excess of that shown by, for example, palmitic acid. Shorter chained fatty acids than those represented in the deposit were less lethal.We wish to thank Dr A. T. James for the chromatographic analyses and for advice; Dr R. J. W. Rees for carrying out counts of total acid-fast bacilli in two experiments; Messrs. Robinson &amp; Sons Ltd, Chesterfield, for information on the industrial processes involved in the preparation of cotton wool; Mr E. J. Latham for advice on preparing glassware; and Dr J. Marks for helpful criticism of the manuscript.