Entomogenous Fungi as Promising Biopesticides for Tick Control

When ticks were sealed in nylon tetrapacks and infected with the entomogenous fungi, Beauveria bassiana and Metarizium anisopliae and maintained in potted grass in the field, the fungal oil formulations (10⁹ conidia per ml) induced 100% mortality in larvae of Rhipicephalus appendiculatus and Amblyomma variegatum, whereas mortalities in nymphs varied between 80–100% and in adults 80–90%. The aqueous formulations (10⁹ conidia per ml) induced mortalities of 40–50% and reductions in egg hatchability of 68% (B. bassiana) and 48% (M. anisopliae) when sprayed on Boophilus decoloratus engorging on cattle. The strains of B. bassiana and M. anisopliae isolated from naturally infected ticks were also found to induce high mortalities in both R. appendiculatus and A.variegatum in tetrapacks placed in potted grass. Both aqueous and oil-based formulations were found to be effective, although the latter induced higher mortalities. These fungal strains in aqueous formulation (10⁸ conidia per ml) suppressed on-host populations of adult R. appendiculatus by 80% (B. bassiana) and 92% (M. anisopliae) when sprayed on tick-infested grass once per month for a period of 6 months. The feasibility of using entomogenous fungi for tick control in the field is discussed.     

Life history of the tick parasitoid Ixodiphagus hookeri (Hymenoptera: Encyrtidae) in Kenya

We conducted laboratory and field experiments to elucidate the life history of Ixodiphagus hookeri, a parasitoid of the ixodid tick Amblyomma variegatum in Western Kenya. Ixodiphagus hookeri females oviposited in unfed host nymphs as well as engorged nymphs, but rarely in engorged larvae. While I. hookeri developed to adults in engorged nymphs, the eggs laid in unfed nymphs disappeared within 2 days after oviposition. As temperature increased, development time of I. hookeri from oviposition to adult emergence in engorged nymphs decreased from 46 days at 23 °C to 35 days at 28 °C, and their immature survival in engorged nymphs decreased from 67% at 23 °C to 22% at 28 °C. No parasitoid adult emerged from hosts at 30 °C. Individual hosts parasitized by single females produced 42–53 adult wasps, 73% of which were females. As a typical pro-ovigenic species, I. hookeri females had an average of 84 mature eggs at emergence and lived only for a few days. When laboratory-reared, unfed nymphs of A. variegatum were attached to cattle for 4–9 days in subsistence farmers’ fields in Western Kenya, 25% of the engorged nymphs and 4% of the unfed nymphs on cattle were parasitized by I. hookeri, demonstrating that I. hookeri females search for and oviposit in A. variegatum nymphs on cattle. Unlike other strains of I. hookeri that overwinter as eggs in unfed nymphs, I. hookeri could continuously reproduce throughout the year in Western Kenya.     

Evaluation of the fungus Beauveria bassiana (Deuteromycotina: Hyphomycetes), a potential biological control agent of Lutzomyia longipalpis (Diptera, Psychodidae)

Visceral leishmaniasis is a zoonosis whose primary vector in Brazil is the sandfly Lutzomyia longipalpis Lutz & Neiva. Presently, efforts to control the vector have not been effective in reducing the prevalence of disease. A possible alternative to current strategies is the biological control of the vector using entomopathogenic fungi. This study evaluates the effects of the fungus, Beauveria bassiana (Bals.) Vuilleman, in different developmental stages of L. longipalpis. Five concentrations of the fungus were utilized ranging from 10⁴ to 10⁸ conidia/ml, with appropriate controls. The unhatched eggs, larvae and dead adults exposed to B. bassiana were sown to reisolate the fungus. The fungus was subsequently identified by polymerase chain reaction (PCR) and DNA sequencing. Exposure to B. bassiana reduced the number of eggs that hatched by 59% (P < 0.01). The longevity of infected adults was 5 days, significantly lower than that of the negative control which was 7 days (P < 0.001). The longevity of the adult sandfly exposed to the positive chemical (pyrethroid, cypermetherin) control was less than 1 day. The effects of fungal infection on the hatching of eggs laid by infected females were also significant and dose-dependent (P < 0.05). With respect to fungal post-infection growth parameters, only germination and sporulation were significantly higher than the fungi before infection (P < 0.001). The identity of the reisolated fungus was confirmed by automated DNA sequencing post-passage in all insect stages. These data show that B. bassiana has good pathogenic potential, primarily on L. longipalpis larvae and adults. Consequently, the use of this fungus in sandfly control programs has potential in reducing the use of chemical insecticides, resulting in benefits to humans and the environment.     

Pathogenicity, formulation and storage of insect pathogenic hyphomycetous fungi tested against Diabrotica speciosa

Studies were conducted tosearch for fungal strains with potentialpathogenicity against Diabrotica speciosa(Germar) (Coleoptera: Chrysomelidae).Among sixteen fungal isolates screenedthe most virulent was a Beauveria bassiana(Balsamo) Vuillemin isolate (FHD13) thatcaused 70% mortality of D. speciosathird instar larvae. The LC₅₀ value ofB. bassiana isolate FHD13 was3.48 × 10¹⁰ conidia/ml.Different temperatures (4, 17 and 26 °C)and vegetable oils (corn, sunflower and canola)used for storage did not significantly affectviability of conidia. A pathogenicity trialagainst D. speciosa larvae performed withthe corn oil formulation (1 × 10⁸ conidia/mlof oil) caused 65% of mortality.     

Infection of Blissus antillus (Hemiptera: Lygaeidae) Eggs by the Entomopathogenic Fungi Metarhizium anisopliae and Beauveria bassiana

This study determined the pathogenicity and virulence of Beauveria bassiana and Metarhizium anisopliae to eggs of the chinch bug Blissus antillus (Hemiptera: Lygaeidae). Eggs were inoculated under laboratory conditions by immersion in concentrations of 1 × 10⁴ and 5 × 10⁶ conidia/ml. Inoculated eggs were kept under controlled conditions. Evaluations were carried out daily for 20 days. M. anisopliae isolates were highly virulent to eggs, even at 1 × 10⁴ conidia/ml. All B. bassiana isolates tested were considered to be of low virulence or avirulent. The most virulent isolate tested was ESALQ 818 (M. anisopliae), which caused 96.7% infection, when eggs were immersed in suspensions of 1 × 10⁴ conidia/ml. Conidial production on infected eggs was observed to be highest for M. anisopliae isolate CG144, with a mean value of 11.6 × 10⁵ conidia/ml/egg. Infection of Blissus eggs oviposited on plant stems was greater when M. anisopliae isolate CG144 was formulated in mineral oil (63.5% mortality) than when formulated in Tween 80 (27.1% mortality).     

The effects of culture media, solid substrates, and relative humidity on growth, sporulation and conidial discharge of Valdensinia heterodoxa

Valdensinia heterodoxa (Sclerotiniacae) is a potential fungal bioherbicide for control of salal (Gaultheria shallon). The effect of culture media, substrates and relative humidity (RH) on growth, sporulation and conidial discharge of V. heterodoxa was determined for two isolates PFC2761 and PFC3027 in vitro. Culture media significantly affected the growth, sporulation, and conidial discharge of V. heterodoxa. Of eight agar media used, colony radial growth was optimal on salal oatmeal agar and salal potato dextrose agar for isolates PFC2761 and PFC3027, respectively; whereas sporulation was at an optimum on salal oatmeal agar for both isolates. Of the eight liquid media tested, mycelial production was highest on wheat bran–salal–potato dextrose broth. Growth on solid substrates greatly stimulated sporulation and conidial discharge of V. heterodoxa. Of the 12 solid substrates used, the greatest numbers of discharged conidia were observed from wheat bran and wheat bran–salal within 14 d of sporulation. Sporulation on solid substrates continued for 42 d. RH significantly affected the sporulation and conidial discharge for both isolates across all solid substrates tested. No conidia were produced or discharged below 93 % RH on wheat bran–salal and millet. With an increase of the RH from 93 to 97 %, sporulation and the number of discharged conidia increased significantly for both isolates on wheat bran–salal, but not on millet.     

Field and laboratory studies on the timing of oviposition and hatching of the western black-legged tick, Ixodes pacificus (Acari: Ixodidae)

The timing of oviposition and hatching of Ixodes pacificus was investigated in the field and at constant temperatures in the laboratory. Replete females held at temperatures between 9 and 29°C began depositing eggs a mean of 9–70 days after drop off. Egg masses held between 12 and 25°C commenced hatching 25–178 days after the onset of oviposition. Eggs held at 9 or 29°C did not hatch. The lower temperature thresholds for development (LTD) for oviposition and hatching were 6.5 and 9°C, respectively. The number of degree days required for oviposition and hatching was 173 and 588, respectively. Replete females placed in the field on 2 December through to 8 March deposited eggs from 2 February through to 24 April; the eggs commenced hatching between 2 July and 21 August. Unfed larvae from two of 20 egg masses survived through the winter and fed readily when exposed to deer mice (Peromyscus maniculatus) on 22 April. Replete larvae were returned to the field and moulted between 9 and 21 August. Larvae exposed to deer mice in August, 4 weeks after hatching, also fed readily. Although further studies are needed to clarify the timing of nymphal development, the present study suggests that I. pacificus requires more than 1 year to complete its life cycle.     

Evaluation of the Entomopathogenic Fungi Beauveria bassiana and Paecilomyces fumosoroseus for Microbial Control of the Silverleaf Whitefly, Bemisia argentifolii

Collaborative research was conducted at the USDA-ARS Subtropical Agricultural Research Center in southern Texas to assess the microbial control potential of Beauveria bassiana and Paecilomyces fumosoroseus against Bemisia whiteflies. Laboratory assays demonstrated the capacity of both pathogens to infect Bemisia argentifolii nymphs on excised hibiscus leaves incubated at relative humidities as low as 25% at 23 ± 2°C (ca. 35% infection by B. bassiana and P. fumosoroseus resulted from applications of 0.6–1.4 × 10³ conidia/mm² of leaf surface). In small-scale field trials using portable air-assist sprayers, applications at a high rate of 5 × 10¹³ conidia in 180 liters water/ha produced conidial densities of ca. 1–2.5 × 10³ conidia/mm² on the lower surfaces of cucurbit leaves. Multiple applications of one isolate of P. fumosoroseus and four isolates of B. bassiana made at this rate at 4- to 5-day intervals provided >90% control of large (third- and fourth-instar) nymphs on cucumbers and cantaloupe melons. The same rate applied at 7-day intervals also provided >90% control in zucchini squash, and a one-fourth rate (1.25 × 10¹³ conidia/ha) applied at 4- to 5-day intervals reduced numbers of large nymphs by >85% in cantaloupe melons. In contrast to the high efficacy of the fungal applications against nymphs, effects against adult whiteflies were minimal. The results indicated that both B. bassiana and P. fumosoroseus have strong potential for microbial control of nymphal whiteflies infesting cucurbit crops.     

Development of a submerged-liquid sporulation medium for the potential smartweed bioherbicide Septoria polygonorum

Submerged culture experiments were conducted in three phases to determine the optimal medium for rapidly producing conidia of the fungal bioherbicide Septoria polygonorum. In phase I, 47 crude carbon sources were evaluated to determine which would support sporulation. Under the conditions tested, pea brine (5–10% v/v) provided best conidiation. In phase II, a fractional factorial design was utilized to screen 38 different medium adjuncts in combination with pea brine for improved sporulation. MgSO₄ was the only factor that resulted in a significant improvement. In phase III, a central composite design with response surface methodology was used to optimize concentrations of these critical factors. The model predicted optimal sporulation in a medium composed of 8.88% v/v pea brine+0.1 molar MgSO₄ with an expected titer of 1.78×10⁸ conidia/ml. Actual mean titer attained with the model-derived medium was 1.15×10⁸ conidia/ml. No significant difference was observed in virulence of conidia produced on agar vs. the model-derived (liquid) medium.     

Induction of diapausing amictic eggs in Synchaeta pectinata

Amictic females of a clone of S. pectinata from Star Lake (Norwich, Vermont) may produce diapausing as well as non-diapausing (subitaneous) eggs. The proportion of diapausing eggs produced in cultures was unaffected by temperature (12 vs 19 °C) or rotifer population density (minima of 0.33 vs 3 ind. ml^–1) at 19 °C. However, at 19 °C this proportion was higher in cultures maintained at a low food level suppressing reproduction (5 × 10³ cells ml^–1 Cryptomonas erosa) than in those maintained at a high food level (2 × 10⁴ cells ml^–1); the treatment effect was marginally significant (p=0.067). Consistent with the effect of low food availability, a period of starvation was very effective in inducing the development of diapausing eggs. None of 19 females cultured individually from hatching at 19 °C on C. erosa (2 × 10⁴ cells ml^–1) in 1-ml volumes produced any diapausing eggs in 4 days (0 out of 349 eggs), while 13 out of 16 females subjected to a 15-hour starvation period 6 hours after birth produced one or more diapausing eggs during that time (34% of the 158 eggs produced by the 16 females were diapausing). Diapausing eggs produced and left at 19 °C hatched after 4 to 13 days. Those produced in cultures with a low food level took significantly longer to hatch (9.7 days) than those produced in cultures with a high food level (8.1 days) (p=0.022). In natural communities, S. pectinata should be able to respond directly and rapidly to poor food conditions by producing eggs that undergo an obligatory dormant period before resuming development.     

Biology of <Emphasis Type="Italic">Dermacentor silvarum</Emphasis> (Acari: Ixodidae) under laboratory conditions

The minimum life cycle of Dermacentor silvarum Olenev had a mean duration of 87.5 days (range 74–102 days) under laboratory conditions [(27±1 °C), 70% RH, 6 L: 18 D]. The mean time in (days) for the different stages of its cycle was as follows: incubation period of eggs was 15.3 days; prefeeding, feeding and premoulting periods of larvae and nymphs averaged 5.5, 4.0 and 7.3 days, and 5.2, 5.0 and 14.6 days, respectively; prefeeding, feeding, preoviposition and oviposition periods of female adults lasted for 7.8, 4.5, 4.3 and 14.0 days, respectively. There existed a highly significant correlation between engorged body weight of females and egg masses laid (r = 0.9877, p<0.001). The reproductive efficiency index (REI) and reproductive fitness index (RFI) in females were 11.09 and 9.58, respectively. No relationship between nymphal engorged body weight and resultant sexes was observed. Delayed feeding and non-oviposition (in June and July) existed in females, and low temperature (−10 °C) treatment for 45 days could terminate oviposition diapause. However, the egg masses laid by post-diapause females were significantly smaller than those laid by females engorged in March, April and May.     

Selection of Beauveria bassiana isolates for control of the whiteflies Bemisia tabaci and Trialeurodes vaporariorum on the basis of their virulence, thermal requirements, and toxicogenic activity

As part of a 3-fold approach to select potential mycoinsecticides for whitefly control, we evaluated infectivity, thermal requirements, and toxicogenic activity of the entomopathogenic fungus Beauveria bassiana (Ascomycota: Clavicipitaceae) under laboratory conditions. Twenty-five native B. bassiana isolates and a commercially available mycoinsecticide (based on B. bassiana) were evaluated for virulence to fourth instar nymphs of sweetpotato whitefly, Bemisia tabaci, and greenhouse whitefly, Trialeurodes vaporariorum, at a concentration of 1 × 10⁷ conidia/ml. All isolates were pathogenic for both whitefly species, whereas mortality rates varied from 3 to 85%. A second series of bioassays was conducted on 10 selected isolates using four 10-fold concentrations ranging from 1 × 10⁵ to 1 × 10⁸ conidia/ml. Median lethal concentrations (LC₅₀) of the four most virulent isolates varied from 1.1 × 10⁵ to 6.2 × 10⁶ conidia/ml and average survival time (AST) of treated nymphs from 5.9 to 7.4 days. T. vaporariorum were significantly more susceptible to all B. bassiana isolates than B. tabaci. The thermal biology of the eight most virulent isolates to both whitefly species was investigated at six temperatures (10–35 °C). The colony radial growth rate was estimated from the slope of the linear regression of colony radius on time and data were then fitted to a modified generalized β function that accounted for 90.5–99.3% of the data variance. Optimum temperatures for extension rate ranged from 23.1 to 27.1 °C, whereas maximum temperatures for fungal growth varied from 31.8 to 36.6 °C. On the basis of their virulence and thermal requirements, three isolates showed promise as candidates for whitefly management in Mediterranean greenhouses. Whilst in vitro production of macromolecular compounds toxic to Galleria mellonella larvae was not a requisite for virulence, ASTs of larvae injected with Sephadex G-25 fractions from candidate isolates ranged from 1.4 to 3.7 days compared with 5–6 days for non-toxic G-25 fractions. In addition, proteinase K treatment significantly reduced their toxic activity suggesting that they were proteins and revealing the potential of these isolates to be further improved through biotechnology to kill the pest more quickly.     

<Emphasis Type="Italic">Beauveria bassiana</Emphasis> pathogenicity to the cherry slugworm, <Emphasis Type="Italic">Caliroa cerasi</Emphasis> (Hymenoptera: Tenthredinidae) larvae

The cherry slugworm Caliroa cerasi is a significant destructive pest of sour cherries (Prunus cerasus) in Turkey. The potential of entomopathogenic fungi for controlling C. cerasi was evaluated. The effects of exposure methods and conidial concentrations (1 × 10⁶, 1.5 × 10⁶, 1 × 10⁷ and 1.5 × 10⁷ conidia/ml) on mature larvae of C. cerasi infected by Beauveria bassiana were investigated under laboratory conditions. Larvae sprayed directly with B. bassiana conidial suspensions and larvae exposed to B. bassiana-treated leaves resulted in 100% mortality within 2.90 and 2.77 days, respectively. The median lethal time (LT₅₀) and days to mortality were highest in the 1.0 × 10⁷ concentrations of conidia for both direct spray and leaf exposure. The present study suggests that B. bassiana has good potential for control of the cherry slugworm, C. cerasi.     

Cold-hardiness of Dermacentor marginatus (Acari: Ixodidae)

The cold-hardiness of Dermacentor marginatus using laboratory-reared offspring of ticks collected in Germany was characterized. Investigations of unfed stages revealed that adult ticks suffered 50% mortality at –10°C after 4–5 months, but larvae and nymphs suffered mortality within few days, whereas –15°C was lethal for all stages within a very short period. Larval hatch and moulting of engorged larvae and nymphs did not occur at 10°C. Embryonic development of eggs with larval hatch was considerably reduced by exposure of eggs to 10°C. Engorged females did not lay eggs at 10°C, the oviposition capability, however, persisted over 6 months at 10°C, 5 months at 5°C, 3 months at 0°C and 2 months at –10°C without substantial decrease of the oviposition capacity or reduction of viable eggs. These results present evidence that unfed adult ticks are the ecoepidemiologically most effective stages, which are capable to tolerate long and extremely cold winters without substantial impairment of the population density. It is also considered that engorged females interrupt their oviposition at low and subzero temperatures delaying it for months and so contribute in bypassing winter conditions. None of the stages survived supercooling indicating that D. marginatus is freeze intolerant. Mean supercooling point (SCP) ranged between –26°C in eggs and –12, 6°C in engorged females. Compared with eggs, the SCP of the other stages was significantly higher. In conclusion, the SCP is considered to have no predictive value in the context with cold-hardiness.     

Development of a submerged-liquid sporulation medium for the johnsongrass bioherbicide<Emphasis Type="Italic"> Gloeocercospora sorghi</Emphasis>

Submerged culture experiments were conducted in three phases to determine the optimal medium for rapidly producing conidia of the fungal bioherbicide Gloeocercospora sorghi. In phase I, 18 crude carbon sources were evaluated to determine which would support sporulation. Under the conditions tested, butter bean and lima bean brines (1.5–4.6 mS/cm) provided best conidiation. In phase II, a fractional-factorial design was utilized to screen 76 different medium adjuncts in combination with butter bean brine for improved sporulation. d-Mannitol and carboxymethylcellulose (CMC) were the only acceptable factors that resulted in a significant improvement. In phase III, a central composite design with response surface methodology was used to optimize concentrations of these critical factors. The model predicted optimal sporulation in a medium composed of 2.69 mS/cm butter bean brine +0.043 M d-mannitol +0.37% w/v CMC with an expected titer of 1.51×10⁷ conidia/ml. Actual mean titer attained with the model-derived medium was 1.91×10⁷ conidia/ml. Optimal sporulation occurred at 25.5°C in this medium and conidia remained viable up to 2.71 days when stored at 12°C. No significant difference was observed in virulence of conidia produced on agar vs washed conidia produced in the model-derived (liquid) medium.     

Biology of Ixodes (Pholeoixodes) hexagonus under laboratory conditions Part II. Effect of mating on feeding and fecundity of females

The effect of mating on the feeding and fecundity ofIxodes (Pholeoixodes) hexagonus females was studied under controlled laboratory conditions of 22–23°C and 98% relative humidity. The feeding period of mated females was 6–15 days and 11–13 days for unmated females. The mean weight of the engorged mated females was 114.84±45.89 mg, whereas, that of the engorged unmated females was significantly lower (80.61±28.84 mg). During the initial slow feeding period, the weight of mated females increased 6.6 times. At the end of the blood feeding, they had increased their initial weight 35.5 times. Unmated females never entered the rapid engorgement phase and up to 12 days of feeding period their mean weight did not increase more than 9.2 times. The pre-oviposition periods of mated and unmated females were 6–15 days and 4–12 days, respectively. The mean of the egg production efficiency was 40.26±12.47% for mated females and 35.68±12.2% for unmated females. The mean of the mass conversion efficiency was 73.6±13.7% for mated females and 66.48 ±16.55% for unmated females. Sixty per cent of the eggs deposited by mated females hatched whereas only 1% of the eggs deposited by unmated females hatched. These results indicate thatI. hexagonus females possess some predisposition for parthenogenesis and only fertility and not fecundity depends on mating.     

Isolation of Beauveria species from Lebanon and evaluation of its efficacy against the cedar web-spinning sawfly, Cephalcia tannourinensis

Larvae of the cedar web-spinning sawfly, Cephalcia tannourinensis Chevin (Hymenoptera: Pamphiliidae), infected with a white fungus were collected from the Tannourine-Hadath El-Jebbeh cedar forest. Macro- and micro-morphological data based on the examination of colonies, conidiophores, and conidial shape of the fungus suggested a Beauveria species. Sequence analysis of the internal transcribed spacer regions of the isolated fungus showed that it is most closely related to isolates of B. bassiana Clade C. The present study showed that the isolated B. bassiana is a naturally occurring entomopathogenic fungus parasitizing the larvae of C. tannourinensis in Lebanon. Laboratory bioassays showed that B. bassiana caused high mortality of eggs and larvae. The infected eggs turned brownish in color, while larvae of the first instar ceased feeding and showed immobility and rigidity within 5 days and before sporulating conidial mat appeared on their cuticle. Second and third larval instars took longer time to show fungal sporulation: mortalities ranged between 85 and 100% within 7 days when treated with different conidial concentrations. The efficacy of control of C. tannourinensis using B. bassiana was higher or equal to the reference insect growth regulator, diflubenzuron, suggesting the possibility of its success as a biological control agent.     

Life-cycle and host specificity of Amblyomma tigrinum (Acari: Ixodidae) under laboratory conditions

Biological data of three generations of Amblyomma tigrinum in the laboratory are reported and the suitability of different host species for immature ticks are compared. Grouping the three generations, infestations by both the larval and nymphal stages were performed on chickens (Gallus gallus), wistar rats (Rattus norvegicus), rabbits (Oryctolagus cuniculus),wild mice (Calomys callosus), dogs (Canis familiaris) and opossums (Didelphis albiventris). Only dogs were used for infestations by adult ticks. Tick developmental periods were observed in an incubator at 27°C and RH 90%. The proportion of engorged larvae recovered from chickens (21.7% of the exposed larvae) was significantly larger (p<0.001) than those from the five mammal species used in the infestations (maximum of 3.1%). A significant larger (p<0.01) proportion of engorged larvae successfully molted after being fed on chickens than on mammal hosts. The proportion of engorged nymphs recovered from chickens (28.8% of the exposed nymphs) was significantly larger (p<0.001) than those from mammal hosts (range: 0–2.1%). Larvae showed similar feeding periods on exposure to different host species, except for those larvae fed on C. callosus, which showed significantly longer (p<0.001) feeding periods. Engorged larvae detachment peaked on the 5^th feeding day, followed by the 6^th day, on all hosts except for C. callosus. Larval premolt periods were similar for engorged ticks exposed to different host species, except for larvae fed on dogs, which showed significantly longer (p<0.001) premolt periods. Host detachment of engorged nymphs peaked on the 6^th feeding day on chickens. Although nymphal detachment on rats peaked on the 8^th day, only 15 nymphs were recovered from this host species. In a sample of 144 F₃ nymphs fed on chickens no significant difference (p>0.10) was found between the feeding or premolt periods of 82 males and 62 females, but female nymphs were significantly heavier (p<0.005) than male nymphs. Sixteen engorged females (61.5% of the exposed ticks) were recovered after being fed on dogs, and all these females laid viable eggs. Chickens, the only avian host, were the most suitable host when compared with the five mammal species. Dogs were demonstrated to be a suitable host for adults of A. tigrinum, which is consistent with, several reports of adult A. tigrinum ticks parasitizing dogs in different areas of South America. Our results reinforce that in these same areas avian species are the major hosts for immature stages of this tick species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Induction of Tetraploid Gynogenesis in the European Sea Bass (Dicentrarchus labrax L.)

A preliminary study on tetraploid gynogenetic induction in the European sea bass was performed by pressure-blocking the second polar body release and the first cleavage in eggs fertilized with ultraviolet-irradiated sperm. Fertilization of eggs with genetically inactivated sperm produced only haploid development that terminated around hatching. Pressure treatments (8.500 psi for 2 min) applied at 6 and 65 min after fertilization (a.f.) produced variable levels (7–95%) of tetraploid larvae at hatching. A small proportion of mosaics (3.8n/4.2n) was also recorded.