Virulence of the entomopathogenic fungus <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> using soybean oil formulation for control of the cotton stainer bug, <Emphasis Type="Italic">Dysdercus peruvianus</Emphasis>

The cotton stainer bug Dysdercus peruvianus (Hemiptera: Pyrrhocoridae) is an insect pest that causes heavy losses in cotton plantations. The need to reduce the use of insecticides for control of this pest has increased steadily, and Metarhizium anisopliae (Ascomycota: Clavicipitaceae) could be an important biopesticide candidate to control this pest. The effect of M. anisopliae on D. peruvianus nymphs and adults using formulations with soybean oil and Agral^® was evaluated. Formulation using 10% soybean oil added to 10⁸ conidia mL^?1 (grown on used and reused rice) was the most effective for nymph and adult, causing 100% mortality 6 and 7 days after exposure, respectively. The SEM analysis of infected insects showed that M. anisopliae conidia were able to adhere anywhere on the exoskeleton, but were more abundant between the joints. Using the same rice for two batches of growth may be an option for improving commercial conidial production of M. anisopliae and may reduce overall costs. Its effect on D. peruvianus adults opens a new possibility for using this fungus as an alternative to chemical pesticides and the use of M. anisopliae in association with integrate pest management.     

Comparative study between Larval Packet Test and Larval Immersion Test to assess the effect of <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> on <Emphasis Type="Italic">Rhipicephalus microplus</Emphasis> tick larvae

Entomopathogenic fungi, such as Metarhizium anisopliae, for the control of arthropods, have been studied for more than 20 years. The aim of this study was to determine the best methodology to evaluate the in vitro effect of the fungus M. anisopliae on Rhipicephalus microplus tick larvae. We compared a modified Larval Packet Test (LPT) and a Larval Immersion Test (LIT). For the LPT filter papers were impregnated with 1 mL of M. anisopliae suspension in Triton X-100 at 0.02%, in concentrations of 10⁶, 10⁷ and 10⁸ conidia/mL and subsequently folded to include the larval ticks. LIT was performed by immersing the larvae in M. anisopliae suspensions for 5 min using the same three concentrations, then the larvae were placed on filter paper clips. For LPT, the LT₅₀ values obtained were 134.6, 27.2 and 24.8 days for concentrations of 10⁶, 10⁷ and 10⁸ conidia/mL; and the mortality after 21 days was 17.3, 17.6 and 38%, respectively. The LT₅₀ values of LIT were 24.5, 20 and 9.2 days with mortality after 21 days of 50.5, 64.7 and 98% for 10⁶, 10⁷ and 10⁸ conidia/mL, respectively. For the same conidia concentration, LIT showed a higher mortality in a shorter time interval when compared with LPT. These differences between the methods tested must be taking into account in further screening and effect studies with M. anisopliae. The set of results shown here could optimize the protocol used to identify M. anisopliae strains pathogenic against R. microplus.     

Complete chloroplast genome sequence of <Emphasis Type="Italic">Capsicum</Emphasis><Emphasis Type="Italic">baccatum</Emphasis> var. <Emphasis Type="Italic">baccatum</Emphasis>

Molecular markers derived from the complete chloroplast genome can provide effective tools for species identification and phylogenetic resolution. Complete chloroplast (cp) genome sequences of Capsicum species have been reported. We herein report the complete chloroplast genome sequence of Capsicum baccatum var. baccatum, a wild Capsicum species. The total length of the chloroplast genome is 157,145 bp with 37.7 % overall GC content. One pair of inverted repeats, 25,910 bp in length, was separated by a small single-copy region (17,974 bp) and large single-copy region (87,351 bp). This region contains 86 protein-coding genes, 30 tRNA genes, 4 rRNA genes, and 11 genes contain one or two introns. Pair-wise alignments of chloroplast genome were performed for genome-wide comparison. Analysis revealed a total of 134 simple sequence repeat (SSR) motifs and 282 insertions or deletions variants in the C. baccatum var. baccatum cp genome. The types and abundances of repeat units in Capsicum species were relatively conserved, and these loci could be used in future studies to investigate and conserve the genetic diversity of the Capsicum species.     

Association between <Emphasis Type="Italic">cagA</Emphasis>, <Emphasis Type="Italic">vacAi</Emphasis>, and <Emphasis Type="Italic">dupA</Emphasis> genes of <Emphasis Type="Italic">Helicobacter pylori</Emphasis> and gastroduodenal pathologies in Chilean patients

In addition to the already known cagA gene, novel genetic markers have been associated with Helicobacter pylori (H. pylori) virulence: the dupA and vacAi genes. These genes might play an important role as specific markers to determine the clinical outcome of the disease, especially the vacAi gene, which has been expected to be a good marker of severe pathologies like gastric adenocarcinoma. In the present study, the association of cagA, dupA, and vacAi genes with gastroduodenal pathologies in Chilean patients was studied. One hundred and thirty-two patients positive for H. pylori were divided into two groups—non-severe and severe gastric pathologies—and investigated for the presence of cagA, dupA, and vacAi H. pylori virulence genes by PCR. The cagA gene was detected in 20/132 patients (15.2%), the vacAi1 gene was detected in 54/132 patients (40.9%), the vacAi2 gene was detected in 26/132 patients (19.7%), and the dupA gene was detected in 50/132 (37.9%) patients. Logistic regression model analysis showed that the vacAi1 isoform gene in the infected strains and the severity of the diseases outcome were highly associated, causing severe gastric damage that may lead to gastric cancer (p < 0.0001; OR = 8.75; 95% CI 3.54–21.64). Conversely, cagA (p = 0.3507; OR = 1.62; 95% CI 0.59–4.45) and vacAi2 (p = 0.0114; OR = 3.09; 95% CI 1.26–7.60) genes were not associated with damage, while the dupA gene was associated significantly with non-severe clinical outcome (p = 0.0032; OR = 0.25; 95% CI 0.09–0.65). In addition, dupA gene exerts protection against severe gastric pathologies induced by vacAi1 by delaying the outcome of the disease by approximately 20 years.     

<Emphasis Type="Italic">Plectosphaerella cucumerina</Emphasis> as a bioherbicide for <Emphasis Type="Italic">Cirsium arvense</Emphasis>: proof of concept

Plectosphaerella cucumerina (Lindf.) W. Gams was evaluated as a bioherbicide for Cirsium arvense L. (Scop.) using a Canadian and a New Zealand isolate. Both isolates defoliated C. arvense when applied at 10¹³ conidia ha^?1 in water volumes ranging from 250 to 6400 l ha^?1 with a rapid decline in effect with declining conidial dose. Repeat application and the addition of the adjuvant Pulse^® penetrant to the conidial suspension increased the disease severity in C. arvense. Maximum disease occurred at 20 °C with a 48 h post-application dew period. The experiments demonstrate that P. cucumerina can defoliate C. arvense under the environmental conditions of temperate pastures where the weed is problematic. The results also show that modifications to formulation and strategic application may reduce the 48 h dew period requirement and risk to non-target species respectively, supporting the conclusion that the fungus has potential as a bioherbicide for C. arvense.     

<Emphasis Type="Italic">Talaromyces heiheensis</Emphasis> and <Emphasis Type="Italic">T. mangshanicus</Emphasis>, two new species from China

Two new species, Talaromyces heiheensis from rotten wood and T. mangshanicus isolated from soil, are illustrated and described as new to science in sections Trachyspermi and Talaromyces. The phylogenetic positions of the two new species inferred from the internal transcribed spacer, beta-tubulin, calmodulin and RNA polymerase II second largest subunit regions were carried out. Talaromyces heiheensis is phylogenetically closely related to T. albobiverticillius, T. rubrifaciens, T. solicola and T. erythromellis, and characterised by slow growth on Czapek yeast autolysate agar at 25 °C, orange conidia en masse on malt extract agar at 25 °C, biverticillate and terverticillate conidiophores, acerose phialides and subglobose to ellipsoidal, smooth-walled conidia. Talaromyces mangshanicus is related to T. kendrickii, T. qii and T. thailandensis, and characterised by slow-growing colonies with absent or sparse sporulation on CYA agar at 25 °C, conidia en masse greyish purple, purplish red soluble pigment on yeast extract agar (YES) at 25 °C, biverticillate conidiophores, ampulliform phialides and subglobose to ellipsoidal conidia with echinulate walls. They are distinguished from the known species in culture characteristics on four standard media, microscopic features and sequence data.     

Susceptibility of <Emphasis Type="Italic">Diaphorina citri</Emphasis> (Hemiptera: Liviidae) and Its Parasitoid <Emphasis Type="Italic">Tamarixia radiata</Emphasis> (Hymenoptera: Eulophidae) to Entomopathogenic Fungi under Laboratory Conditions

Diaphorina citri (Kuwayama) is a global pest of citrus that transmits the bacteria associated with the disease, Huanglongbing. Entomopathogenic fungi and the parasitoid Tamarixia radiata (Waterston) are important biological control agents of this pest and likely to interact in D. citri populations. As a basis for interaction studies, we determined the susceptibility of nymphs and adults of D. citri and adults of the parasitoid T. radiata to six fungal isolates from the species Beauveria bassiana s.l. (Bals.-Criv.) Vuill. (isolates B1 and B3), Metarhizium anisopliae s.s. (Metsch.) (Ma129 and Ma65) and Isaria fumosorosea Wize (I2 and Pae). We conducted experiments evaluating infection levels in all three insect groups following inoculation with a series of conidial concentrations (1 × 10⁴–1 × 10⁸ conidia mL^?1). Results showed that D. citri nymphs and T. radiata were more susceptible to fungal isolates than D. citri adults. Overall, B. bassiana and M. anisopliae isolates caused the greatest infection compared with I. fumosorosea isolates in all three groups of insects. Isolates B1 (B. bassiana) and Ma129 (M. anisopliae) infected a greater proportion of adults and nymphs of D. citri, respectively. Both isolates of B. bassiana caused greater infection in T. radiata compared with isolates of the other fungal species. We propose that isolates B1 and Ma129 are the strongest candidates for control of D. citri. Our results represent the first report of entomopathogenic fungi infecting T. radiata, and the basis for future studies to design a biological control programme that uses both agents more efficiently against D. citri populations.     

Phylogenetic assessment and taxonomic revision of <Emphasis Type="Italic">Mariannaea</Emphasis>

Four new species of Mariannaea were described in this paper, namely M. chlamydospora, M. cinerea, M. fusiformis, and M. lignicola. Mariannaea chlamydospora is characterized by its cream-colored, zonate colonies on PDA, smooth conidiophores, fusiform conidia, and abundant chlamydospores. Mariannaea cinerea forms grey colonies and ellipsoidal to subglobose conidia. Mariannaea fusiformis forms purple colonies and fusiform to subglobose conidia. Mariannaea lignicola has brown conidiophores and broad hyphae. The molecular phylogeny was inferred using ITS, LSU, and TUB-2 loci. The type species of Mariannaea (M. elegans) is epitypified. The variety M. elegans var. punicea is raised to species rank. Mariannaea clavispora is excluded from Mariannaea because of its cylindrical phialides, straight conidial chains and deviating phylogenetic affinity. Mariannaea nipponica did not fit well the generic concept of Mariannaea based on their morphological characters, and its generic placement remains uncertain. A key to the currently accepted 15 species of Mariannaea is provided.     

Histological and cytological studies of plant infection by <Emphasis Type="Italic">Erysiphe euonymi</Emphasis>-<Emphasis Type="Italic">japonici</Emphasis>

Powdery mildew caused by Erysiphe euonymi-japonici (Eej) is an increasingly serious fungal disease on Euonymus japonicus that is an important ornamental plant. However, little is currently known about infection and pathogenesis of Eej on E. japonicus. Here, we report plant infection by Eej at the histological and cytological levels. Eej caused severe disease symptoms with white and snow-like colonies on leaf surfaces of E. japonicus. Microscopic observations were conducted continuously to define infection process of Eej on E. japonicus. Eej conidia germinated to produce appressorial germ tubes on leaf surfaces and formed irregular haustoria in plant epidermal cells at 6 h post-inoculation (hpi) and 12 hpi, respectively. After uptaking nutrients from host cells by haustoria, Eej formed numerous hyphae and extensive colonization on leaf surfaces at 96 hpi and finally produced abundant conidiophores and new conidia on leaf surfaces at 168 hpi. In addition, there was consistently a single nucleus in different Eej infection structures and haustorial development could be divided into three major stages, including formation of penetration peg, formation of haustorial neck and initial haustorium, and maturation of haustorium. These results provide useful information for further determination of Eej pathogenesis and finally controlling the disease.     

<Emphasis Type="Italic">Aeromonas</Emphasis> and <Emphasis Type="Italic">Plesiomonas</Emphasis> species from scarlet ibis (<Emphasis Type="Italic">Eudocimus ruber</Emphasis>) and their environment: monitoring antimicrobial susceptibility and virulence

The present study aimed at evaluating the role of captive scarlet ibises (Eudocimus ruber) and their environment as reservoirs of Aeromonas spp. and Plesiomonas spp., and analyzing the in vitro antimicrobial susceptibility and virulence of the recovered bacterial isolates. Thus, non-lactose and weak-lactose fermenting, oxidase positive Gram-negative bacilli were recovered from cloacal samples (n = 30) of scarlet ibises kept in a conservational facility and from water samples (n = 30) from their environment. Then, the antimicrobial susceptibility, hemolytic activity and biofilm production of the recovered Aeromonas spp. and Plesiomonas shigelloides strains were assessed. In addition, the virulence-associated genes of Aeromonas spp. were detected. Ten Aeromonas veronii bv. sobria, 2 Aeromonas hydrophila complex and 10 P. shigelloides were recovered. Intermediate susceptibility to piperacillin-tazobactam and cefepime was observed in 2 Aeromonas spp. and 1 P. shigelloides, respectively, and resistance to gentamicin was observed in 4 P. shigelloides. The automated susceptibility analysis revealed resistance to piperacillin-tazobactam and meropenem among Aeromonas spp. and intermediate susceptibility to gentamicin among P. shigelloides. All Aeromonas isolates presented hemolytic activity, while 3 P. shigelloides were non-hemolytic. All Aeromonas spp. and 3/10 P. shigelloides were biofilm-producers, at 28 °C, while 10 Aeromonas spp. and 6/10 P. shigelloides produced biofilms, at 37 °C. The most prevalent virulence genes of Aeromonas spp. were asa1 and ascV. Scarlet ibises and their environment harbour potentially pathogenic bacteria, thus requiring monitoring and measures to prevent contamination of humans and other animals.     

Effects of adjuvant and conidial concentration on the efficacy of <Emphasis Type="Italic">Beauveria bassiana</Emphasis> for the control of the two spotted spider mite, <Emphasis Type="Italic">Tetranychus urticae</Emphasis>

Greenhouse experiments were conducted on various crops (cucumber, tomato, eggplant, green bean) to ascertain the effects of Break-thru^® (polyether-polymethylsiloxane-copolymer, a silicone surfactant) and an oil emulsion, on Beauveria bassiana (Balsamo) Vuillemin (Bb) applications for the control of the two spotted spider mite, Tetranychus urticae Koch. The objectives were to compare a) the efficacy of Bb control when applied in aqueous Break-thru^® or an oil emulsion; b) the effects of various concentrations of Bb conidia, as affected by each surfactant; and c) the effects of Break-thru^® on the activity of the fungus. Conidia were suspended either in an aqueous Break-thru^® or an emulsifiable formulation at different conidial concentrations (1.05 × 10⁶, 2.1 × 10⁶ and 4.2 × 10⁶ conidia ml^?1) and sprayed onto leaves 2 weeks after artificial pest inoculation. Two sprays were performed, with an interval of one week from one spray to another, and T. urticae population counts (both motile and egg stages) were made on plant leaves 7 days after each spray. Bb conidia in Break-thru^® were more efficacious than conidia in emulsifiable formulation. With the highest rate of conidia (4.2 × 10⁶ conidia ml^?1), mortality of adult mites ranged from 60 ± 4.2 (mean ± SE) to 85.7 ± 4.3% in the Break-thru^® suspension and 39.4 ± 7 to 61.3 ± 6% in the oil emulsion. Leaf damage index was also substantially reduced from 70% in the unsprayed control to 40% by the application of Bb conidia at the highest rate with Break-thru^®. Break-thru^® can be combined with Bb in the integrated management of T. urticae and Isolate R444 is a promising candidate for the control of the pest.     

In vitro efficacy of <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> sensu lato against unfed <Emphasis Type="Italic">Amblyomma parvum</Emphasis> (Acari: Ixodidae)

Amblyomma parvum Aragão (Acari: Ixodidae) is a tick species found with wide distribution in the Neotropical region. Even though it is a wildlife-related tick, it is also a frequent parasite of domestic animals, is aggressive to human beings and may harbor pathogenic microorganisms. Therefore, it is a target species for control on domestic animals, particularly those at the rural–wildlife interface. Herein, the efficacy of two isolates (E9 and IBCB 425) of an entomopathogenic fungus, Metarhizium anisopliae sensu lato, already evaluated for ticks that parasitize domestic animals, was tested against unfed A. parvum adults. Both isolates displayed high acaricidal efficacy after immersion in fungal conidial suspensions for 5 min. Isolate E9 killed all ticks by the 7th day post-treatment, and isolate IBCB 425 did so by the 11th day. Tick mortality of 80 and 90% was achieved as early as the 3rd and 4th days, respectively, with both treatments. Thus, if a commercial M. anisopliae s.l. acaricide against domestic animal ticks is developed, it would also be effective against A. parvum.     

<Emphasis Type="Italic">R</Emphasis>-acetoin accumulation and dissimilation in <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis>

Klebsiella pneumoniae is a 2,3-butanediol producer, and R-acetoin is an intermediate of 2,3-butanediol production. R-acetoin accumulation and dissimilation in K. pneumoniae was studied here. A budC mutant, which has lost 2,3-butanediol dehydrogenase activity, accumulated high levels of R-acetoin in culture broth. However, after glucose was exhausted, the accumulated R-acetoin could be reused by the cells as a carbon source. Acetoin dehydrogenase enzyme system, encoded by acoABCD, was responsible for R-acetoin dissimilation. acoABCD mutants lost the ability to grow on acetoin as the sole carbon source, and the acetoin accumulated could not be dissimilated. However, in the presence of another carbon source, the acetoin accumulated in broth of acoABCD mutants was converted to 2,3-butanediol. Parameters of R-acetoin production by budC mutants were optimized in batch culture. Aerobic culture and mildly acidic conditions (pH 6–6.5) favored R-acetoin accumulation. At the optimized conditions, in fed-batch fermentation, 62.3 g/L R-acetoin was produced by budC and acoABCD double mutant in 57 h culture, with an optical purity of 98.0 %, and a substrate conversion ratio of 28.7 %.     

Usefulness of the Non-conventional <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis> Model to Assess <Emphasis Type="Italic">Candida</Emphasis> Virulence

Invasive candidiasis is caused mainly by Candida albicans, but other Candida species have increasing etiologies. These species show different virulence and susceptibility levels to antifungal drugs. The aims of this study were to evaluate the usefulness of the non-conventional model Caenorhabditis elegans to assess the in vivo virulence of seven different Candida species and to compare the virulence in vivo with the in vitro production of proteinases and phospholipases, hemolytic activity and biofilm development capacity. One culture collection strain of each of seven Candida species (C. albicans, Candida dubliniensis, Candida glabrata, Candida krusei, Candida metapsilosis, Candida orthopsilosis and Candida parapsilosis) was studied. A double mutant C. elegans AU37 strain (glp-4;sek-1) was infected with Candida by ingestion, and the analysis of nematode survival was performed in liquid medium every 24 h until 120 h. Candida establishes a persistent lethal infection in the C. elegans intestinal tract. C. albicans and C. krusei were the most pathogenic species, whereas C. dubliniensis infection showed the lowest mortality. C. albicans was the only species with phospholipase activity, was the greatest producer of aspartyl proteinase and had a higher hemolytic activity. C. albicans and C. krusei caused higher mortality than the rest of the Candida species studied in the C. elegans model of candidiasis.     

Characterization and virulence of <Emphasis Type="Italic">Lecanicillium lecanii</Emphasis> against different aphid species

Seven isolates of Lecanicillium lecanii (Zimmermann) Zare &; Gams isolated in Spain from infected aphids were characterized using sequences of the Internal Transcribed Spacer (ITS) regions and also based on morphological and physiological characteristics. Four of these seven L. lecanii isolates were selected to assess their virulence against nymphs of Myzus persicae (Sulzer), Nasonovia ribisnigri (Mosley), Macrosiphum euphorbiae (Thomas) and Aphis gossypii Glover. Mortality (%), lethal concentration 50 (LC₅₀) and lethal time 50 (TC₅₀) were calculated. The analysis of the sequences of ITS region confirmed that the new isolates were clearly Lecanicillium lecanii. The set of isolates had similar radial growth (51.5–54.0 mm), except for ICAL1 (39 mm). The germination time 50 (GT₅₀) varied between 10.7 h (ICAL3) and 13 h (ICAL5). The isolate ICAL6 showed the highest value for conidial production (3.4 × 10⁸ con ml^?1) and also produced the highest mortality for M. persicae (95%) and was more virulent than the commercial product Vertalec^® (91.6%).     

Betulin inhibits cariogenic properties of <Emphasis Type="Italic">Streptococcus mutans</Emphasis> by targeting <Emphasis Type="Italic">vicRK</Emphasis> and <Emphasis Type="Italic">gtf</Emphasis> genes

Streptococcus mutans, a multivirulent pathogen is considered the primary etiological agent in dental caries. Development of antibiotic resistance in the pathogen has created a need for novel antagonistic agents which can control the virulence of the organism and reduce resistance development. The present study demonstrates the in vitro anti-virulence potential of betulin (lup-20(29)-ene-3β,28-diol), an abundantly available plant triterpenoid against S. mutans UA159. Betulin exhibited significant dose dependent antibiofilm activity without affecting bacterial viability. At 240 µg/ml (biofilm inhibitory concentration), betulin inhibited biofilm formation and adherence to smooth glass surfaces by 93 and 71 % respectively. It reduced water insoluble glucan synthesis by 89 %, in conjunction with down regulation of gtfBC genes. Microscopic analysis confirmed the disruption in biofilm architecture and decreased exopolysaccharide production. Acidogenicity and aciduricity, key virulence factors responsible for carious lesions, were also notably affected. The induced auto-aggregation of cells upon treatment could be due to the down regulation of vicK. Results of gene expression analysis demonstrated significant down-regulation of virulence genes upon betulin treatment. Furthermore, the nontoxic effect of betulin on peripheral blood mononuclear cells even after 72 h treatment makes it a strong candidate for assessing its suitability to be used as a therapeutic agent.     

Production of <Emphasis Type="Italic">trans</Emphasis>-10,<Emphasis Type="Italic">cis</Emphasis>-12-conjugated linoleic acid using permeabilized whole-cell biocatalyst of <Emphasis Type="Italic">Yarrowia lipolytica</Emphasis>

Objective

To improve the production of trans-10,cis-12-conjugated linoleic acid (t10,c12-CLA) from linoleic acid in recombinant Yarrowia lipolytica.

Results

Cells of the yeast were permeabilized by freeze/thawing. The optimal conditions for t10,c12-CLA production by the permeabilized cells were at 28 °C, pH 7, 200 rpm with 1.5 g sodium acetate l^?1, 100 g wet cells l^?1, and 25 g LA l^?1. Under these conditions, the permeabilized cells produced 15.6 g t10,c12-CLA l^?1 after 40 h, with a conversion yield of 62 %. The permeabilized cells could be used repeatedly for three cycles, with the t10,c12-CLA extracellular production remaining above 10 g l^?1.

Conclusion

Synthesis of t10,c12-CLA was achieved using a novel method, and the production reported in this work is the highest value reported to date.

     

The <Emphasis Type="Italic">yajC</Emphasis> gene from <Emphasis Type="Italic">Lactobacillus buchneri</Emphasis> and <Emphasis Type="Italic">Escherichia coli</Emphasis> and its role in ethanol tolerance

The yajC gene (Lbuc_0921) from Lactobacillus buchneri NRRL B-30929 was identified from previous proteomics analyses in response to ethanol treatment. The YajC protein expression was increased by 15-fold in response to 10 % ethanol vs 0 % ethanol. The yajC gene encodes the smaller subunit of the preprotein translocase complex, which interacts with membrane protein SecD and SecF to coordinate protein transport and secretion across cytoplasmic membrane in Escherichia coli. The YajC protein was linked to sensitivity to growth temperatures in E. coli, involved in translocation of virulence factors during Listeria infection, and stimulating a T cell-mediated response of Brucella abortus. In this study, the L. buchneri yajC gene was over-expressed in E. coli. The strain carrying pET28byajC that produces YajC after isopropyl β-d-1-thiogalactopyranoside induction showed tolerance to 4 % ethanol in growth media, compared to the control carrying pET28b. This is the first report linking YajC to ethanol stress and tolerance.     

Distribution and habitats of <Emphasis Type="Italic">Phengaris</Emphasis> (<Emphasis Type="Italic">Maculinea</Emphasis>) butterflies and population ecology of <Emphasis Type="Italic">Phengaris</Emphasis><Emphasis Type="Italic">teleius</Emphasis> in China

Many species of the butterfly genus Phengaris are regarded as endangered in many parts of their distribution. Several species are also widely distributed across northern China. Due to land use change and overgrazing, their habitats are declining and many patches have been lost. This paper investigates the distribution and habitats of the Chinese Phengaris species (of the subgenus Maculinea). Shrub-grassland near forests seem the most frequent habitat for Phengaris, while flat open grasslands are mostly over-grazed and thus survival for Phengaris butterflies there seems difficult. Throughout Europe, P. teleius is an endangered species, while there is still no information on its status in China. To improve the knowledge on the population ecology of P. teleius, its population structure, adult behaviour and movement were studied through mark–release–recapture methods in the Qinling Mountains of Taibai County. Eight grassland patches which were potentially suitable were found in the area in 2013. In total, 480 individuals (274 females) were marked, resulting in an overall recapture rate of 16 %. The average daily population size was 44 butterflies (±23 SD) during the adult flight period. Sixty-seven percent of the females and 38 % of the males moved less than 50 m, and 17 % of recaptured females and 38 % of males moved more than 200 m. The mean movement distance was 107 ± 177 m for males and 182 ± 122 m for females. The majority of the recaptures (86 %) were made within the patches, only a few individuals (14 %) moved between patches. Due to human disturbance and destruction, all of the eight potentially suitable patches are becoming smaller and increasingly isolated, thus these populations of P. teleius may face an increasing risk of extinction, which may well be a tip of the iceberg of habitat loss and fragmentation of P. teleius in Taibai County and possibly beyond. Hence we hope our initial study of P. teleius could have positive impacts on the conservation of Phengaris butterflies in China.