On the Nature of nontypable Haemophilus influenzae

193 Haemophilus cultures, including 71 nontypable H. influenzae isolates, were examined with respect to phage HP1 sensitivity, lysogeny for this and for other phages and for excretion of bacteriocins. Fifty of the 71 nontypable cultures were sensitive to phage HP1 but only three produced plaques. The other 47 isolates were thus probably not non-encapsulated derivatives of H. influenzae serotypes a, b, d, and e, which have discrete and characteristic phage HP1 restriction and modification systems, or serotype c which appears to be restriction negative. They could be derivatives of serotype f which does not give plaques with phage HP1. The nontypable three cultures that plated phage HP1 efficiently could be non-encapsulated serotype c derivatives. Fourteen of the phage HP1 insentitive nontypable cultures were found to be defectively lysogenic for this phage. Five of these were genetically transformed to wild type lysogens. Their phage produced plaques efficiently only on Rc strains and on a restriction-negative mutant of serotype d. These lysogenic nontypable isolates are thus modification (and restriction) negative and they are thus probably not nonencapsulated derivatives of serotypes a, b, d, e, or f. Fifty three of 56 serotype b cultures were found to excrete a bacteriocin, to which all other nonproducing Haemophilus cultures were more or less sensitive. The three restriction-negative nontypable H. influenzae cultures also excreted this bacteriocin but the other cultures listed did not do this. The tentative conclusion from this study is that nontypable H. influenzae isolates are probably not derivatives of the six known encapsulated strains.     

Isolation and Characterization of Actinomyces propionicus

Three cultures of Actinomyces have been identified as Actinomyces propionicus. Two of these strains are recent isolates, one, 427, from a case of cervico-facial actinomycosis, and one, 439, from a case of lacrimal canaliculitis. The third strain, 346, was described by F. Lentze as A. israelii serological type II. These three strains were compared with the type strain of A. propionicus ATTC 14157 and with known strains of five other Actinomyces species. Morphologically and biochemically the three new cultures of A. propionicus were identical with the type strain but closely resembled A. israelii. In serological tests making use of fluorescent antibody, all four A. propionicus strains gave negative results with antisera for A. israelii, A. bovis, A. naeslundii, and A. eriksonii, but gave positive results with antisera for A. propionicus 14157 and strain 346. The A. propionicus antisera did not stain other Actinomyces species. A. propionicus contains diaminopimelic acid (DAP) in its cell wall and produces propionic acid from glucose. All three new isolates were shown to contain DAP and to produce propionic acid. By use of the presence of DAP in the cell wall and serological tests as the differential criteria, the three cultures described in the report were specifically identified as A. propionicus.     

Reisolation of Staphylococcus salivarius from the human oral cavity

Twenty-four strains of gram-positive facultative cocci, arranged primarily in small clusters, were isolated from the surface of the human tongue. With the exception of 14 catalase-negative isolates, these strains were identical in cultural and biochemical characteristics and in deoxyribonucleic acid base composition. All cultures produced viscous growth in both liquid and agar media. They fermented glucose anaerobically, reduced nitrate beyond nitrite, were benzidine-positive, and failed to grow in the presence of 5% NaCl or at 45 C. In addition, they exhibited guanine plus cytosine (G + C) contents of 55.4 to 58.3%. These isolates differed from strains of pediococci, aerococci, and micrococci which were included for comparison. On the basis of G + C content, these organisms appear to be intermediate between micrococci and staphylococci; however, on the basis of anaerobic glucose fermentation, it is suggested that they be placed in the genus Staphylococcus. It is proposed that they be recognized as S. salivarius.     

Occurrence and characterization of "avium-like" Mycobacteria isolated from animals in Sweden

431 cultures of "avium-like" mycobacteria (ALM) were isolated from wild and domestic animals during 1974--76 at the National Veterinary Institute, Stockholm. Of these, 50 isolates from pigs were studied by growth-chromogenicity, pathogenicity, and biochemical tests. Furthermore, thin-layer chromathography was performed, and on some isolates serotyping. All 50 isolates belonged to Runyon's group III and were pathogenic for chicken; none was capable of splitting oleic acid from Tween 80. 47 gave tellurite reduction within a period of three days; one was arylsulphatase-positive after three days and a further four after 14 days. The biological and biochemical tests permit assignation of the 50 isolates to the M. avium-intracellulare complex. The lipid patterns of the isolates examined were analysed by thin-layer chromatography. Thirty-five of the isolates showed a lipid pattern similar to that of A 2 of the fowl reference strain; three belonged to lipid type A 1 and four to A 3. Eight could not be typed. Of 22 isolates, 14 could be assigned to M. avium serotypes.     

Distribution of plasmid- and chromosome-mediated iron uptake systems in Vibrio anguillarum strains of different origins

We investigated the incidence of plasmid-mediated and chromosome-mediated iron uptake systems in strains of Vibrio anguillarum that belong to serotypes O1 and O2 and were isolated from different fish species and in different geographic areas. All of the strains gave positive reactions in CAS agar medium and in the Arnow test, which indicated that catechol types of siderophores were produced. The majority of V. anguillarum serotype O1 strains harbored a 65-kb plasmid similar to plasmid pJM1 from strain 775, which encodes the siderophore anguibactin and its outer membrane receptor, protein OM2. All of the isolates harboring this plasmid promoted the growth of an anguibactin-deficient receptor-proficient mutant derived from strain 775, but none of these isolates promoted the growth of mutants lacking receptor OM2. Furthermore, under iron-limiting conditions all of these strains induced outer membrane proteins that were identical in size to protein OM2 of strain 775. In contrast, none of the serotype O2 strains contained a high-molecular-weight plasmid, but all of them induced the growth of mutants defective in the anguibactin-mediated system regardless of the presence or absence of receptor OM2. The serotype O2 strains, but not the plasmid-bearing serotype O1 strains, also induced the growth of Salmonella typhimurium enb-1 which utilizes only enterobactin as a siderophore.(ABSTRACT TRUNCATED AT 250 WORDS)     

Distribution of plasmid- and chromosome-mediated iron uptake systems in Vibrio anguillarum strains of different origins.

We investigated the incidence of plasmid-mediated and chromosome-mediated iron uptake systems in strains of Vibrio anguillarum that belong to serotypes O1 and O2 and were isolated from different fish species and in different geographic areas. All of the strains gave positive reactions in CAS agar medium and in the Arnow test, which indicated that catechol types of siderophores were produced. The majority of V. anguillarum serotype O1 strains harbored a 65-kb plasmid similar to plasmid pJM1 from strain 775, which encodes the siderophore anguibactin and its outer membrane receptor, protein OM2. All of the isolates harboring this plasmid promoted the growth of an anguibactin-deficient receptor-proficient mutant derived from strain 775, but none of these isolates promoted the growth of mutants lacking receptor OM2. Furthermore, under iron-limiting conditions all of these strains induced outer membrane proteins that were identical in size to protein OM2 of strain 775. In contrast, none of the serotype O2 strains contained a high-molecular-weight plasmid, but all of them induced the growth of mutants defective in the anguibactin-mediated system regardless of the presence or absence of receptor OM2. The serotype O2 strains, but not the plasmid-bearing serotype O1 strains, also induced the growth of Salmonella typhimurium enb-1 which utilizes only enterobactin as a siderophore.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation of cellulolytic mesophilic clostridia from a municipal solid waste digestor

Ten obligately anaerobic, cellulolytic mesophilic bacteria were isolated from a municipal solid waste digestor used for biogas production. The isolates were rod-shaped, spore-forming bacteria in anaerobic conditions, and stained Gram-positive in young cultures, and hence were identified asClostridium. Small regular translucent and unpigmented colonies were observed on cellulose plates. The strains were gelatinase-negative, hydrolyzed esculin and starch, and fermented xylose and arabinose. The lecithinase, lipase, and indole tests were negative. The major fermentation products from cellulose included ethanol and acetate. The morphological and other biochemical characteristics indicated that these clostridia did not correspond to any previously described species. All the strains produced high activities of extracellular cellulases in cellulose media and degraded paper. Offprint requests to: L. Benoit.     

Antigenicity of a viral peptide displayed on β-galactosidase fusion proteins is influenced by the presence of the homologous partner protein

Abstract In an earlier study of the distribution of O-serotypes among clinical isolates of Serratia marcescens , two apparently new serotypes were identified, represented by strains S1254 and S3255. Studies using ELISA, immunoblotting and the Quellung reaction have shown that they qualify for inclusion in the O-antigenic typing scheme on three counts: (1) they possess chemically distinct O-antigenic repeating units, (2) the O-antigens are serologically distinguishable from all others, and (3) they are found in a significant proportion of clinical S. marcescens strains (13% and 6% respectively). S1254, the type strain for serotype O27, is an acapsular strain which expressed a glucorhamnan with a disaccharide repeating unit as its lipopolysaccharide side chain. It cross-reacts with serotype O4, the O antigen of which is an O-acetylated form of the O27 glucorhamnan, but this cross-reaction can be eliminated by reciprocal cross-absorption. S3255, the type strain for serotype O28, has a mannose homopolymer as its O-antigen and is the only S. marcescens serotype with a trimeric repeating-unit structure. However, it cross-reacts with the O5 serotype strain due to similarities in their acidic capsular polysaccharides. Cross-absorption and the production of serum to an acapsular variant of serotype strain O28 produced typing reagents which could differentiate serotypes O5 and O28.     

Ecological studies on the occurrence of bacteria utilizing lactic acid at pH values below 4.5

Sites within two cheese factories, a silage tower and silage pit and two pet food factories were investigated for lactic acid utilizing bacteria. The bacteria isolated, together with known lactic acid utilizing and aciduric bacteria, were tested for their ability to 'alkalinize' a fermented meat product containing more than 20 g/1 lactic acid at pH > 4.3 a ten-fold dilution of this product and synthetic media, both at pH 4.5. Strains able to alkalinize the diluted product were 13 aerobic isolates (all Acetobacter spp.), nine anaerobic cultures, one strain of Megasphaera elsdenii and two strains of Clostridium tyrobutyricum. Eight of the aerobes could alkalinize undiluted product containing < 1 g/1 residual potassium sorbate, but had no effect on product containing > 3 g/1 sorbate; none of the anaerobic cultures was able to alkalinize undiluted product. Acetobacter spp., therefore, threatened the microbial stability of a fermented product containing < 3 g/1 potassium sorbate. The cultures able to alkalinize diluted product were all from sites contaminated with whey, silage or fermented product for at least 2 d, suggesting substantial accumulation of lactic acid utilizing bacteria at these sites.     

Population Snapshot of Streptococcus pneumoniae Causing Invasive Disease in South Africa Prior to Introduction of Pneumococcal Conjugate Vaccines

We determined the sequence types of isolates that caused invasive pneumococcal disease (IPD) prior to routine use of pneumococcal conjugate vaccines (PCV) in South Africa. PCV-13 serotypes and 6C isolates collected in 2007 (1 461/2 437, 60%) from patients of all ages as part of on-going, national, laboratory-based surveillance for IPD, were selected for genetic characterization. In addition, all 134 non-PCV isolates from children <2 years were selected for characterization. Sequence type diversity by serotype and age category (children <5 years vs. individuals ≥5 years) was assessed for PCV serotypes using Simpson’s index of diversity. Similar genotypes circulated among isolates from children and adults and the majority of serotypes were heterogeneous. While globally disseminated clones were common among some serotypes (e.g., serotype 1 [clonal complex (CC) 217, 98% of all serotype 1] and 14 [CC230, 43%)]), some were represented mainly by clonal complexes rarely reported elsewhere (e.g., serotype 3 [CC458, 60%] and 19A [CC2062, 83%]). In children <2 years, serotype 15B and 8 were the most common serotypes among non-PCV isolates (16% [22/134] and 15% [20/134] isolates, respectively). Sequence type 7052 and 53 were most common among serotypes 15B and 8 isolates and accounted for 58% (7/12) and 64% (9/14) of the isolates, respectively. Serotype 19F, 14, 19A and 15B had the highest proportions of penicillin non-susceptible isolates. Genotypes rarely reported in other parts of the world but common among some of our serotypes highlight the importance of our data as these genotypes may emerge post PCV introduction.     

Two new subspecies of Bacillus thuringiensis isolated in Japan: Bacillus thuringiensis subsp. kumamotoensis (serotype 18) and Bacillus thuringiensis subsp. tochigiensis (serotype 19)

Bacteriological and serological characteristics of three Bacillus thuringiensis isolates obtained in Japan were investigated. They formed typical rhomboidal parasporal inclusions but flagellar (H) antigens of these isolates were different from those of the known 17 H serotypes of B. thuringiensis. The three isolates were divided into two new serotypes (serotypes 18 and 19). The serotype 18 isolate (3–71) produced thermostable exotoxin and the inclusions of this isolate were toxic to larvae of the silkworm, Bombyx mori, but nontoxic to larvae of the mosquito, Aedes aegypti. The other isolate (119-72) belonging to serotype 18 produced inclusions nontoxic to larvae of B. mori and A. aegypti and did not produce thermostable exotoxin. However, other bacteriological properties of the isolate 119-72 were similar to those of the isolate 3–71. The serotype 19 isolate (117-72) produced inclusions nontoxic to larvae of B. mori and A. aegypti and did not produce thermostable exotoxin. Acid production from saccharose and the production of brownish purple pigment were observed in the two serotype 18 isolates, while neither of them was observed in the serotype 19 isolate. In other 29 biochemical properties tested, there was no difference among the three isolates. Based on these characteristics, the following two subspecies names are proposed: Bacillus thuringiensis subsp. kumamotoensis (serotype 18) for the type strain 3–71 and Bacillus thuringiensis subsp. tochigiensis (serotype 19) for the type strain 117-72.     

Lipopolysaccharide heterogeneity in Pasteurella haemolytica isolates from cattle and sheep.

Lipopolysaccharide (LPS) from 40 isolates of Pasteurella haemolytica, comprising 23 serotype A1, seven serotype A2, one serotype T4, one serotype T10 and eight untypable isolates, obtained from diseased and healthy cattle or sheep, was characterized by SDS-PAGE and Western blotting. Ten different SDS-PAGE LPS profiles, five smooth and five rough, were identified among the biotype A and untypable isolates and designated LPS types 1-10. LPS types 1 and 2 were smooth, had similar O-antigen banding-patterns but differed in the low-molecular-mass or core-oligosaccharide regions; type 3 LPS was rough but had a core-oligosaccharide region similar to that of LPS type 1. No similarities were observed between these LPS types and types 6, 7 and 9, which were smooth, and types 4, 5, 8 and 10, which were rough. Most serotype A1 isolates (19/23) were of LPS type 1, whereas two isolates each had LPS of types 2 and 3. The majority (5/7) of serotype A2 isolates possessed type 3 LPS, whereas the remaining two isolates each had LPS of types 4 and 5. There was much greater heterogeneity within the untypable group of isolates, which comprised LPS of types 1 and 9 (two isolates each), and 6, 7, 8 or 10 (one isolate each). Western blotting analysis demonstrated that LPS types 1 and 2 had immunologically identical O-antigen side-chains but differed in their core-oligosaccharide regions, whereas the core-oligosaccharide region of rough LPS type 3 was immunologically very similar to that of LPS type 1. The other LPS types were immunologically unrelated to these three LPS types. The majority (20/23) of serotype A1 isolates originated from cattle and possessed LPS types 1 or 2, different from most (5/7) of the serotype A2 isolates which originated from sheep and possessed LPS of types 3 or 4. However, two of the three ovine serotype A1 isolates had the same type 3 LPS as occurred in most of the ovine serotype A2 isolates, suggesting a possible correlation between LPS type and host specificity. This study has demonstrated that LPS diversity within different serotypes of P. haemolytica is greater than was previously thought and that certain LPS types might be host-specific.     

Identification of Actinomyces israelii and Actinomyces naeslundii by Fluorescent-Antibody and Agar-Gel Diffusion Techniques

This study was an attempt to develop a fluorescent-antibody (FA) test to differentiate Actinomyces israelii and A. naeslundii as an aid in their laboratory identification. Two strains of A. israelii (X522 and A601) and two strains of A. naeslundii (X454 and X600), which had received intensive study by several investigators, were used for the immunization of rabbits. Working titers, based on tests with antigens prepared from the homologous strains and from well-established heterologous strains, were determined for each labeled antibody preparation. These conjugates and their normal serum control conjugates were used separately to stain 85 cultures of Actinomyes species and 23 strains of other species that might be confused with them. Acetone-precipitated soluble antigens from these same strains were tested with different antisera in the agar-gel diffusion test. Results showed that A. israelii (X522 and A601) and A. naeslundii (X454 and X600) labeled antiglobulins, when used at their working titers, stained most strains of their homologous species. Agar-gel diffusion results showed general agreement with those of the FA tests. The two tests appear to be equal in sensitivity, but the FA test is more specific, since several cross-reactions were noted with the agar-gel diffusion test whereas no cross-reactions were obtained with the FA reagents. Agar-gel and FA studies suggest that at least two serotypes of A. israelii may be associated with human disease. Although the majority of strains tested in this study appear to belong to a common serotype, "serotype 1," two strains of an apparent second serotype, "serotype 2," were encountered. FA staining of tissue impression smears from experimentally infected mice was successful when a counterstain, Evans Blue dye, was used.     

Identification of virulence factors in Vibrio cholerae isolated from Iraq during the 2007-2009 outbreak

Thousands of people were infected with Vibrio cholerae during the outbreak in Iraq in 2007-2009. Vibrio cholerae was shown to be variable in its content of virulence determinants and in its antibiotic sensitivity. This study was designed to isolate and characterize clinical and environmental V.?cholerae isolates and to determine antibiotic sensitivity, enzyme and toxin production, and the presence of virulence genes. Eighty clinical and five environmental bacterial isolates were collected and diagnosed by subjecting them to microscopic, biochemical, serological, and molecular analysis. The results revealed that 55% of clinical isolates belonged to the Inaba serotype, 32.5% to the Ogawa serotypes, and 12.5% to the Non-O1 serotype. All environmental V.?cholerae isolates belonged to the Non-O1 serotype. All environmental isolates were sensitive to all examined antimicrobial agents, while all clinical isolates showed a high sensitivity (100%) to ampicillin, gentamicin, cephalothin, tetracycline, erythromycin, and ciprofloxacin, and a high resistance (97.5%) to co-trimoxazole, nalidixic acid, and chloramphenicol. It was found that all V.?cholerae (O1) isolates were resistant to the Vibrio static O129 and all Non-O1 V.?cholerae isolates were sensitive to the Vibrio static O129. All clinical and environmental isolates produced hemolysin (100%) and lecithinase (100%), while they showed various production rates of protease (90% of clinical and 60% of environmental) and lipase (50% of clinical and 20% of environmental). The ompW gene was amplified in all the clinical and environmental V.?cholerae isolates, but not in other related and nonrelated bacteria. Multiplex PCR analysis showed that the toxR gene was amplified in all clinical and environmental isolates, while ctxA, ctxB, tcpA genes were amplified only in clinical (O1) isolates. This study indicates the differences in the production of some enzymes and toxins and in the content of virulence genes between clinical and environmental isolates in Iraq during the outbreak (2007-2009).     

Electrophoretic karyotypes of <Emphasis Type="Italic">C. neoformans</Emphasis> serotype A recovered from Thai Patients with AIDS

Thirty-seven clinical isolates of C. neoformans were recovered from AIDS patients and all were serotype A according to standard typing tests. They were further analyzed using RAPD, PCR fingerprinting, and PFGE along with 2 additional reference isolates ATCC 34871 (serotype A) and RV 45981 (serotype D). Using 2 different RAPD primers, all of the clinical isolates and the reference serotype A (ATCC 34871) gave similar RAPD patterns while serotype D (RV 45981) gave distinctive pattern. Corresponding result was also obtained upon PCR by using a primer for microsatellite (GACA)₄. However, using a primer specific to minisatellite M13 + 1, all PCR fingerprinting gave similar gel patterns (M1) for 35/37 of the clinical isolates and the reference serotype A while two clinical isolates generated different patterns called M2 and M3. The reference serotype D gave distinctive pattern called M4. PFGE gave 17 different karyotypes that could be categorized into 4 groups named EKA (1–6), EKB (1–5), EKC (1– 5) and EKD (1). The reference serotype A fell into group EKA as EKA6 while the reference serotype D fell into group EKC as EKC5. Among the clinical isolates, EKA group (20/37 isolates) and type EKA1 (16/20) dominated with only one isolate each for types EKA2 to EKA5. The next most prevalent was group EKB (12/37 isolates) which dominately fell in type EKB1 (8/12) and only one isolate each for types EKB2 to EKB5. Group EKC (4/37 isolates) and group EKD (1/37) had only one isolate for each type (EKC1 to EKC 4 and EKD1). The 2 predominant karyotypes (EKA1, 16/37 and EKB1, 8/37) may represent two originally common clones of C. neoformans expose among the patients. The high discriminatory power of PFGE infers the benefit of subtyping which lead to better understanding on the epidemiology and pathogenic potential of C. neoformans subtypes. Moreover, PCR fingerprinting and RAPD infer the feasibility of detail analysis between serotypes A and D for unencapsulated C. neoformans.     

Actinomyces georgiae sp. nov., Actinomyces gerencseriae sp. nov., designation of two genospecies of Actinomyces naeslundii, and inclusion of A. naeslundii serotypes II and III and Actinomyces viscosus serotype II in A. naeslundii genospecies 2

DNAs of type strains and representative members of Actinomyces groups from the human periodontal flora and from other habitats were compared by using the S1 nuclease procedure to determine their genetic relatedness. One rather common group from the human periodontal flora, previously called "Actinomyces D08," is phenotypically distinct from, and genetically unrelated to, previously described species. We propose the name of Actinomyces georgiae for this organism; the type strain is strain ATCC 49285. Another common group from the human periodontal flora is Actinomyces israelii serotype II, which was found genetically distinct from the type strain of A. israelii (serotype I) and from other previously described species of Actinomyces. We propose the name Actinomyces gerencseriae for this organism; the type strain is strain ATCC 23860. A. naeslundii serotype I strains were distinct from the other strains studied. A separate genospecies which included strains of A. naeslundii serotypes II and III and A. viscosus serotype II was delineated. Strains of Actinomyces serotype WVA 963 constitute an additional distinct genospecies. Because there are no reliable phenotypic tests, other than serological analyses, to differentiate Actinomyces serotype WVA 963 and the two genospecies of A. naeslundii, no taxonomic changes are proposed for these three genospecies.     

Isolation of Methanobrevibacter smithii from human feces.

Fecal specimens from nine adults were examined for the presence of methanogenic bacteria. Enrichment cultures of five specimens produced methane in 5 days. Of these five specimens, three were tested and produced methane during a short-term incubation. Four specimens did not produce methane in either short-term incubation or in enrichment culture. Each methanogenic culture contained methanogens similar in morphology to organisms of the genus Methanobrevibacter and showed factor-420 fluorescence by fluorescence microscopy. Pure cultures were obtained from four of the five methanogenic enrichment cultures. Each isolate grew and formed methane from either H2-CO2 or formate, but growth obtained with formate was poor. None of the isolates used acetate, methanol, or trimethylamine. All isolates grew in the presence of bile salts. In immunological studies, each isolate was closely related to the type strain of Methanobrevibacter smithii, a finding consistent with the physiological and morphological similarities between the isolates and the type strain.     

Co-existence of colonies with different serotypes and other biological characteristics in clinical isolates of Pseudomonas aeruginosa.

The biological characteristics of individual colonies of Pseudomonas aeruginosa from 138 specimens were investigated. Of these isolates, 90 (65.2%) formed colonies of similar appearance and morphology, and 48 (34.8%) formed colonies which differed either in appearance or morphology. The individual colonies of 138 isolates were tested for serotype. The former 90 isolates formed only the colonies with one kind of serotype, whereas 17 of the latter 48 isolates formed the colonies with more than one kind of serotype. All the 9 isolates tested also differed in other biochemical characteristics: acid productions from xylose, mannitol and maltose, urease production and gelatin liquefaction. beta-Lactamase activity was investigated in 7 isolates forming colonies with more than one serotype. There were no marked differences in beta-lactamase activity among the different colonies in 5 isolates but marked differences among those in the other 2 isolates.     

Effect of nutrition on growth and virulence of the entomopathogenic fungus Beauveria bassiana

Three isolates of the entomopathogen Beauveria bassiana along with one strain of Metarhizium anisopliae were cultured on seven media with different carbon/nitrogen (C/N) ratios. The effect of nutrition on virulence of the isolates was evaluated via measurement of colony growth, spore yield, germination speed, conidial C/N ratio and Pr1 (a serine protease) activity. 'Osmotic stress' medium produced the lowest colony growth with low numbers of conidia in all isolates. However, these conidia showed a high germination rate and virulence. However, conidial Pr1 activity was low in some isolates. In most but not in all cases conidia from 1% yeast extract, 2% peptone and low (10 : 1) C/N medium had higher Pr1 activity compared with conidia from other media. However, in some instances we could not conclude that there was a relationship among germination rate, conidial Pr1 activity and virulence. C/N ratio of conidia was statistically different among various media and fungal isolates. Conidia with lower C/N ratio generally produced lower LT(50) (lowest median lethal time) values (more virulent). Insect-passaged conidia from different media had lower C/N ratio compared with similar conidia from artificial cultures. Therefore, they should be more virulent than in vitro produced conidia. As germination rate, conidial Pr1 activity and C/N ratio are independent of host, it seems that host-related determinants such as insect cuticle and physiology and environmental conditions may influence host susceptibility and therefore fungal isolate virulence towards host insects.