Milky Disease Bacteria

A comparative study was made of all available milky-disease species and strains that have been isolated around the world from beetle larvae (family Scarabaeidae). Included in the study were Bacillus popilliae Dutky, B. lentimorbus Dutky, and B. lentimorbus var. maryland from the United States; B. euloomarahae Beard and B. lentimorbus var. australis Beard from Australia; B. fribourgensis Wille from Switzerland; and New Zealand milky disease (Dumbleton). The organisms were classified into three groups: (i) those containing parasporal bodies, including B. popilliae Dutky, B. fribourgensis Wille, and New Zealand milky disease (Dumbleton); (ii) those without a visible parasporal body and with spore morphology similar to B. lentimorbus Dutky, including B. lentimorbus var. australis Beard; and (iii) those with very tiny spores and no parasporal body, including B. euloomarahae Beard and B. lentimorbus var. maryland. All available milky-disease species and strains were cultivated in vitro on Brain Heart Infusion Agar plates. However, the most fastidious organisms—B. euloomarahae and B. lentimorbus var. maryland—could not be grown until they were passed through a life cycle in larvae of a large scarabaeid beetle infesting rotting wood. Then they remained stable for only one or two subcultures. All the milky-disease organisms produced larger cells in vitro than they did in vivo. The pattern of sugar fermentations was similar for all milky-disease species. It appears that there is a very low percentage of strains of B. popilliae, B. lentimorbus, and the other milky-disease organisms that have the inherent genetic makeup to permit them to sporulate on artificial media, if conditions are favorable. Among these conditions are a sufficiently high cell population and a reduced oxygen tension. Spores produced in vitro may have a low virulence via the normal ingestion pathway, even though they show apparent virulence when injected directly into the hemocoel.     

Role of Leuconostoc mesenteroides in Leavening the Batter of Idli, a Fermented Food of India

The fermentation of the batter of idli, a fermented food of India, was studied. The microorganisms responsible for the characteristic changes in the batter were isolated and identified. Although there is a sequential change in the bacterial flora, the predominant microorganism responsible for souring, as well as for gas production, was found to be Leuconostoc mesenteroides. In the later stages of fermentation, growth of Streptococcus faecalis and, still later, of Pediococcus cerevisiae becomes significant. The fermentation of idli demonstrates a leavening action caused by the activity of the heterofermentative lactic acid bacterium, L. mesenteroides. As far as is known, this is the first record of a leavening action produced exclusively by the activity of a lactic acid bacterium.     

Susceptibility of insecticide-susceptible and wild house flies (Diptera: Muscidae) to abamectin on whitewashed and unpainted wood

Unpainted plywood panels treated with 0.1% abamectin (avermectin B1) provided greater than 90% control of house flies, Musca domestica L., susceptible to insecticides for 4 wk and greater than 70% control for 7 wk compared with 46-92% control observed with permethrin at the same time and rate of application. Efficacy of abamectin on whitewashed panels was similar to that observed on unpainted panels, whereas permethrin was ineffective on whitewashed panels at all rates tested (range, 0.001-0.1%) at all intervals after treatment. Bioassays of newly colonized house flies resistant to permethrin indicated that wild populations may be cross-resistant to abamectin.     

Fermented foods, feeds and beverages

There has been a proliferation of books and papers dealing with the indigenous fermented foods/beverages of the world. It is anticipated that these foods/beverages will play an ever-increasingly important role in feeding both the developing and the developed world as population increases from approximately 4.5 billion to 6 billion by the year 2000 and to 8 to 12 billion people in the 21st century. The indigenous fermented foods consist of microbial protein grown on edible substrates. Microbial or single cell protein (SCP) per se continues to receive research and development attention. It is likely to play an important role in feeding animals in the future when it becomes competitive with soy protein. It may play a direct role in feeding humans in the future after its safety for feeding animals has been adequately demonstrated and it has been shown that it can be processed into foods acceptable to humans. At the present time, mushrooms, a form of microbial protein highly acceptable to humans, which can be grown readily on ligno-cellulosic and other agricultural and food processing wastes, offer considerable opportunity for expanding man's food supply.     

Production of higher alcohols during indonesian tapé ketan fermentation

A study was made of the higher alcohols (fusel oils) produced during the Indonesian tapé ketan fermentation using Amylomyces rouxii as the principal mold, alone or in combination with yeasts belonging to genera commonly found in the tapé ketan fermentation (Endomycopsis, Candida, and Hansenula). Total fusel oils increased with length of fermentation. Fusel oils detected in the product distillate included isobutanol and isoamyl and active amyl alcohols. No n-propanol was detected. Isobutanol and isoamyl alcohols were formed in the largest amounts. A. rouxii alone produced nearly the same quantity of fusel oils (total production, 275 mg/liter at 192 h) as it did in combination with Endomycopsis burtonii (total production, 292 mg/liter at 192 h).A. rouxii and Endomycopsis fibuliger produced fusel oils totaling 72 mg/liter at 32 h and 558 mg/liter at 192 h. A. rouxii in combination with Candida yeasts produced somewhat more fusel oils, ranging from 590 to 618 mg/liter at 192 h. A. rouxii in combination with Hansenula yeasts produced the least fusel oils, totaling 143 to 248 mg/liter at 192 h. During the first 36 h, production of fusel oils was higher at 30 and 35 degrees C than at 25 degrees C. At 48 h fusel oil production was slightly higher at 30 degrees C than at 35 degrees C. Beyond 48 h, production of fusel oils was higher at 25 degrees C. A. rouxii in combination with Hansenula anomala and Hansenula subpelliculosa produced considerable ethyl acetate, ranging from 145 to 199 mg/liter at 36 h and 354 to 369 mg/liter at 192 h.     

Effects of long-term storage at -14 degrees C on the survival of Neozygites fresenii (Entomophthorales: Neozygitaceae) in cotton aphids (Homoptera: Aphididae)

Neozygites fresenii-infected Aphis gossypii cadavers, containing dormant hyphal bodies of N. fresenii, were stored in 4 ml glass vials at -14 degrees C in a standard consumer-type refrigerator/freezer for 1, 21, 30, 43, 51, and 68 months to determine the effect of storage on fungal survival. When the cadavers were removed from the freezer and placed in 25+/-1 degrees C, 100% relative humidity, and 12:12 (L:D) conditions, N. fresenii survival, as shown by fungal sporulation from the cadavers, was high at all storage periods. The average percentage of cadavers from which the fungus sporulated were 93, 47, 100, 100, 80, and 60% from 1, 21, 30, 43, 51, and 68 months storage periods, respectively. The number of primary conidia discharged from each sporulating cadaver was estimated using a scale of 1 (low, ca. 1000 primary conidia), 2 (medium, ca. 2000 primary conidia) and 3 (high, ca. 3000 primary conidia). The median scores for the number of primary conidia produced per sporulating cadaver were 3, 2, 3, 3, 2.5, and 1 for 1, 21, 30, 43, 51, and 68 months, respectively. Therefore, except for the longest storage period, most cadavers produced medium to high numbers of primary conidia. Mean germination of primary conidia produced from N. fresenii-infected-aphid cadavers from each time period varied significantly from 66.3 to 86.1% in the 21 and 43 months categories, respectively. Infectivity of capilliconidia, produced from frozen N. fresenii, to live healthy cotton aphids varied significantly from 16.7 to 68.7% from cadavers stored 68 months and 1 month, respectively. Overall N. fresenii survived well in dried frozen cotton aphid cadavers for up to 6 years with little reduction in sporulation, numbers of spores produced, germination of primary conidia, or infectivity.     

Growth of Paenibacillus larvae,the causative agent of American foulbrood in honey bees and Bacillus popilliae,the causative agent of milky disease in beetle larvae in lepidopteran cell cultures

TNM-FH Lepidopteran insect cell culture medium containing 10% fetal bovine serum (FBS), while allowing limited vegetative growth of Paenibacillus larvae (wild-type strain), the causative agent of American foulbrood, contained no viable vegetative cells upon subculture, nor were any heat resistant spores produced in this medium alone. However, TNM-FH medium cotaining embryonic or midgut cells from Trichoplusia ni, hemocytes from Estigmene acrea, ovarian and embryonic cells from Spodoptera frugiperda, embryonic cells from Plutella xylostella, Spodoptera exigua and Pseudaletia unipuncta or ovarian cells from Lymantria dispar, supported both heavy vegetative cell growth and moderate production of heat resistant spores. EX-CELL 405 serum-free insect cell culture medium alone appeared to contain the appropriate nutrients required for both vegetative growth and sporulation of P. larvae. However, in the presence of embryonic cells from T. ni, limited vegetative growth occurred and the P. larvae cells appeared to die off. This was confirmed by the fact that no colony growth occurred upon subculture, nor were any heat resistant spores detected. This was true also in the presence of fat body cells from T. ni, except that a limited number of spores (4,000/ml) were detected in the form of cology-forming units (CFU) on plates following heating to 80°C for 20 minutes. In a parallel study with a wild-type strain of Bacillus popilliae, vegetative cells grew only in TNM-FH medium in the presence of mid-gut BTI-Tn-MG and ovarian (Tn-368) cells of T. ni. No heat resistant spores, however, were detected in any of the cultures. When BTI-Tn-MG and Tn-368 cells were further challenged with four variant cultures of B. popilliae, vegetative growth and limited sporulation were achieved. The BTI-Tn-MG cell line in TNM-FH medium produced as many as 12,000 spores/ml after 21 days in culture.     

Prevalence of Pandora neoaphidis (Zygomycetes: Entomophthorales) infecting Nasonovia ribisnigri (Hemiptera: Aphididae) on lettuce crops in Argentina

Lettuce crops, Lactuca sativa, organically produced in La Plata, Argentina, were sampled in order to determine the prevalence of fungal diseased aphids. Nasonovia ribisnigri was the only aphid detected and its occurrence was highly variable. The fungal pathogen Pandora neoaphidis (Entomophthoromycotina: Entomophthorales) was the only pathogen detected. We recorded a maximum of 34.2 aphids per plant and the highest rate of fungal prevalence was 56.6% (n = 30) (aphids infected/total aphids). Infected aphids were observed in all sampling sites. No differences of infection rates were detected between the center and the edge of crops. Host density was an important factor determining infection. The majority of host population was comprised of nymphs which were the most infected in terms of individuals per habitat unit (lettuce plant), but considering the proportion of infected aphids per stage of development, the prevalence of infection in nymphs and adults was similar.     

Sensitive assays for simian foamy viruses reveal a high prevalence of infection in commensal, free-ranging Asian monkeys

Foamy viruses (FV) are retroviruses that naturally infect many hosts, including most nonhuman primates (NHPs). Zoonotic infection by primate FV has been documented in people in Asia who reported contact with free-ranging macaques. FV transmission in Asia is a concern, given abundant human-NHP contact, particularly at monkey temples and in urban settings. We have developed three assays capable of detecting the presence of FV in Asian NHP species that are commensal with humans: enzyme-linked immunosorbent assay (ELISA), Western blot assays using recombinant viral Gag protein, and an indicator cell line that can detect macaque FV. The recombinant ELISA correlates very well with the presence of FV sequences detected by PCR. We have used these assays to demonstrate both that FV is highly prevalent among free-ranging NHPs and that seroconversion occurs at a young age in these animals. These assays should also prove useful for large-scale analysis of the prevalence of FV infections in human populations in Asia that are commensal with free-ranging NHPs.