BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, researchlibraries, and research funders in the common goal of maximizing access to critical research.

Life Cycle of Ixodes (Ixodes) loricatus (Acari: Ixodidae) Under LaboratoryConditionsAuthor(s): Teresinha Tizu Sato Schumaker, Marcelo Bahia Labruna, Isis Dos Santos Abel, and PalomaTeixeira Soares ClericiSource: Journal of Medical Entomology, 37(5):714-720. 2000.Published By: Entomological Society of AmericaDOI: http://dx.doi.org/10.1603/0022-2585-37.5.714URL: http://www.bioone.org/doi/full/10.1603/0022-2585-37.5.714

BioOne (www.bioone.org) is a nonprofit, online aggregation of core research in the biological, ecological, andenvironmental sciences. BioOne provides a sustainable online platform for over 170 journals and books publishedby nonprofit societies, associations, museums, institutions, and presses.

Your use of this PDF, the BioOne Web site, and all posted and associated content indicates your acceptance ofBioOne’s Terms of Use, available at www.bioone.org/page/terms_of_use.

Usage of BioOne content is strictly limited to personal, educational, and non-commercial use. Commercial inquiriesor rights and permissions requests should be directed to the individual publisher as copyright holder.

ARTICLE

Life Cycle of Ixodes (Ixodes) loricatus (Acari: Ixodidae) UnderLaboratory Conditions

TERESINHA TIZU SATO SCHUMAKER, MARCELO BAHIA LABRUNA,1 ISIS DOS SANTOS ABEL,AND PALOMA TEIXEIRA SOARES CLERICI

Departamento de Parasitologia, Instituto de Ciencias Biomedicas, Universidade de Sao Paulo, Sao Paulo, SP, Brazil

J. Med. Entomol. 37(5): 714Ð720 (2000)

ABSTRACT The life cycle of Ixodes (Ixodes) loricatus Neumann, reared in the laboratory, isdescribed. Engorged females collected from opossums trapped in the states of Minas Gerais and SaoPaulo, Brazil, which were used to start the laboratory colonies, were designated as BMG and CSP,respectively. Larval and nymphal ticks from both colonies fed separately on Rattus norvergicusBerkenhout or Calomys callosus Rengger, whereas Didelphis marsupialis L and Didelphis albiventrisLund were used as hosts for BMG and CSP adults, respectively. Biological and developmental dataobtained from ticks of both the BMG and CSP colonies that were reared separately for twoconsecutive generations were compared. The percentage of fed or molted ticks reared on C. callosuswas higher than that recorded for ticks fed on R. norvergicus in the majority of the observations.Despite signiÞcant differences among several of the biological parameters, the pattern of the lifecycles of the two tick colonies was similar. Results indicated that the mean life cycle duration of I.(I.) loricatus was '7 mo from parental oviposition to the occurrence of F1 eggs, regardless ofgeographic origin or host species.

KEY WORDS Ixodes (Ixodes) loricatus, life cycle, hosts

DURING THE LAST two decades, several studies havebeen conducted on the biology, epidemiology, andtaxonomic status of the genus Ixodes, as well as theirinteraction with various pathogens. Recent investiga-tions determined the capability of ticks for transmit-ting Borrelia burgdorferi Johnson, Schmid, Hyde,Steigerwalt & Brenner sensu lato to humans or do-mestic animals, indicating tick potential for maintain-ing enzootic foci of borreliosis (Gray 1998, Oliver1999). In the Neotropical Region, where '37 Ixodestick species have been reported (Clifford et al. 1973),the life cycles of only Ixodes (Ixodes) affinis Neumann(Oliver et al. 1987) and Ixodes (I.) minor Neumann(Banks et al. 1998) are known. Both species also arefound in the Nearctic Region.

Ixodes (Ixodes) loricatus Neumann, a common ec-toparasite of opossums, has been reported from Ta-basco de La Frontera in Mexico to Terra del Fuego inArgentina (Nuttal and Warburton 1911, Aragao 1936,Cooley and Khols 1945, Clifford et al. 1973). This tickalso has been reported on numerous rodent and eden-tate species, and even on some primates (Cooley and

Kohls 1945). Despite the wide distribution of I. (I.)loricatus and its preference for opossum species (Di-delphis spp.) that can easily acquire synanthropic hab-its, little biological data for this species has been pub-lished. I. (I.) loricatus seems to transmit borreliaÐlikemicroorganisms among rats and opossums trapped inthe Itapevi andCotia counties in the stateof SaoPaulo,Brazil (Barros-Battesti 1998, Abel 1999). In these geo-graphical areas, humans with clinical symptoms sim-ilar to those of Lyme disease were conÞrmed by se-rological tests (ELISA and Western Blotting) using B.burgdorferi s.s. as the test antigen (Yoshinari et al.1993a, 1993b). InBrazil, the agent of the disease seemsto have an antigenic identity with B. burgdorferi andprobably is responsible for a Lyme disease similis ornew borreliosis (Yoshinari et al. 1999). The aim of thecurrent investigation was to study the life cycle of I.(I.) loricatus under laboratory conditions.

Materials and Methods

Ticks. Ticks used in the study were the progenies ofengorged females collected from Þeld-caught opos-sums. One white-belly opossum, Didelphis albiventrisLund, was captured in Belo Horizonte county(198499 S; 438579 W), state ofMinasGerais, and a singlefemale tick found on it was used to establish a colonydesignated as BMG. One common opossum, DidelphismarsupialisL., was captured inCotia county (238369 S;468559 W), state of Sao Paulo, and a single female tickfound on it was used to establish a colony designated

We followed the protocol which agrees with Ethical Principles inAnimal Research adopted by Brazilian College of Animal Experimen-tation (COBEA), and which was approved by the Biomedical Sci-ences Institute/USP-Ethical Committee for Animal Research(CEEA). Permits and approvals are on Þle in the ofÞce of T.T.S.S.

1 Departamento de Medicina Veterinaria Preventiva e Saude An-imal ÐFaculdade de Medicina Veterinaria e Zootecnia da Univer-sidade de Sao Paulo, Av. Corifeu de Azevedo Marques, 2720, SaoPaulo, SP, Brasil, 05340Ð000.

0022-2585/00/0714Ð0720$02.00/0 q 2000 Entomological Society of America

CSP. The opossums were held in captivity until theticks had fed and detached. Two generations of eachcolony were studied for each stage of tick develop-ment. Throughout the study, the ticks were main-tained at 278C and 95Ð98% RH in an incubation ovenand were reared in total darkness except while tickswere feeding on the host.

Hosts.Weused naiveWistar rats,Rattus norvergicusBerkenhout, and naive wild rodents, Calomys callosusRengger, as hosts for the immature stages of ticks fromboth of the BMG and CSP tick colonies. Each rodentwas placed in a polypropylenebox (30by 20by 13 cm)covered by a wire mesh top and Þlled with a smallquantity of wood shavings. Double-sided adhesivetape was placed around the top edges of the box toprevent tick escape. Opossums D. albiventris and D.marsupialis served as the blood source for adult BMGand CSP ticks, respectively. Four opossums of eachspecies were trapped in the same area where theparent ticks were collected, and were maintained inindividual wire cages (80 by 70 by 60 cm). Each cagewaskeptonaaluminumtable(100by100cm)coveredwith paper towels and surrounded by double-sidedadhesive tape at the edges. Opossums were fed a dietof commercial cat chow ad libitum, and occasionalfruit and eggs and were provided with a daily supplyof water. During tick feeding, the hosts were main-tained at 24Ð278C and 55% RH, and were exposed tonatural daylight cycles.

Feeding. The number of tick feeding days wererecorded from the day after the ticks were put on thehost until the day they were engorged and detachedfrom the host. After the larvae hatched or the nymphsand adults ecdysed, the newly emerged ticks wereheld for a speciÞed prefeeding period of '50, 30 and40 d, respectively, before being placed on a host. Nosystematic attempts were made to determine the min-imumandmaximumdurationof theprefeedingperiodof the three stages; thus, they did not reßect theminimum hardening or maximum survival of eachstage. After being counted, the ticks were brushedonto the dorsal surface of the hosts, using a camelÕs-hair brush. The cage, tray, chow, water, and woodshavings were examined twice daily at 0700 and 1900hours to recover the engorged ticks.

Immature Stages. From the two generations of theBMG tick colony, 3,000 larvae and 410 nymphs wereplaced on eight (375 larvae per host) and 10 (41nymphs per host) R. norvegicus, respectively. Thesame procedures were carried out using C. callosus asthe host. From the two generations of the CSP tickcolony, 2,875 larvae and 255 nymphs were placed oneight (358Ð359 larvae per host) and 10 (25Ð26 nymphsper host) R. norvegicus, respectively. Again, the sameprocedures were used for infesting C. callosus exceptthe total number of nymphs was 335 (33Ð34 per host).

As engorged larvae or nymphs detached, they werecollected and placed in separate vials plugged withcotton, and each individual was observed daily todetermine the time of molting. After nymphal ecdysis,their sex was recorded.

Adults. All males and females that emerged fromimmature ticks fed onR. norvegicus orC. callosuswerekept together in the same screen vial (44 cm3, 3.5 cmdiameter) during all the prefeeding period, but sep-arated by colonies (BMG and CSP), to allow for mat-ing as reported for Prostriata ticks (Oliver 1974, Aub-ert 1981, Farkas and Surgeoner 1991). After 4Ð6 wkthe ticks were allowed to feed on the opossums. Fromthe combined two generations, 51 BMG and 65 CSPadult ticks were placed on four D. albiventris and fourD. marsupialis, respectively. A minimum of six and amaximumof14 femaleandmalepairswere fedoneachindividual host. Engorged females were collected andtheir weights were recorded immediately after de-tachment from the host. Females were observed dailyfor the preoviposition period (the period between theday of detachment and the day when oviposition be-gins) and the oviposition period (the number of daysbetween the beginning and the ending oviposition).Daily deposited eggs were carefully separated fromfemales to verify when the oviposition period ended.

Eggs. The eggs of each female were placed in sep-arate glass tubes (80 by 12 mm) that were pluggedwith cotton and observed daily to determine the num-ber of days from the start of oviposition until larvaewere Þrst observed. After hatching, the percentage ofhatched eggs was determined, always by the sameobserver (Drummond et al. 1973). The mean weightof one egg was calculated from the weight of 10batches of 100 eggs deposited by three separate fe-males per colony.The total numberof eggs in eacheggmass was estimated by dividing the weight of the totalmass of the eggs by the mean weight of one egg. Forthe measurement of the metabolic activity of femalesfor the oviposition process, the index of egg produc-tion efÞciency was determined using the formula:weight of eggs/weight of the engorged female 3 100(Bennett 1974).

Statistical Analysis. Differences between the feed-ingorpremolt periods of larvae andnymphs frombothcolonies (BMGandCSP) fed ondifferent host specieswere determined using two-way analysis of variance(ANOVA) and the Tukey multiple comparison test.The requirements for equality of variances were pre-viously stated and, when necessary, the data weretransformed. Statistical differences between the bio-logical data obtained for BMG and CSP engorgedfemales was determined by using the Student t-testafter the equality of variances was stated. The sametestwasused tocomparebiologicaldataof theÞrst andthe second BMG engorged female generations. Thepercentage of ticks recovered after feeding or moltinganddropping off the host during both night or daywasanalyzed using the chi-square test. The signiÞcantdifference in all tests was P , 0.05.

Results

Immature Stages. The biological and developmen-tal data, excluding the prefeeding periods, obtainedfrom two separate generations of rearing for the BMGand CSP colonies are presented in Table 1.

September 2000 SCHUMAKER ET AL.: LIFE CYCLE OF I. (I.) loricatus 715

Feeding Periods. The feeding period of ticks fromthe same colony fed on different host species showedthat larvae of both colonies fed faster (P , 0.05) on R.norvegicus than those fed on C. callosus (Table 1).However, in both colonies, no signiÞcant differencewas observed between the duration of the feedingperiod of nymphs fed on R. norvegicus compared withthose fedonC. callosus.Analysisof ticks feedingon thesame host species showed that BMG larvae or nymphsreared on R. norvegicus fed faster (P , 0.05) than theCSP ticks (Table 1). Similarly, BMG immature ticksfed faster on C. callosus (P , 0.05) than did CSP ticks.

Premolt Periods. Larvae from both tick coloniesthat fed on R. norvegicus required a signiÞcantlygreater time to undergo ecdysis (P , 0.05) than thosefed on C. callosus (Table 1). No signiÞcant difference(P . 0.05) was detected between the duration of thepremolt period recorded for BMG nymphs fed on R.norvegicus and that of nymphs fed on C.callosus. How-ever, the CSP nymphs fed on R. norvegicus requiredmore time to undergo ecdyses (P , 0.05) than thosefed on C. callosus.

Comparing the duration ofBMG immature tick pre-molt periods to that of theCSP ticks showed thatBMGlarvae fed both on R. norvegicus or on C. callosus

required less time to undergo ecdysis (P , 0.05) thanCSP larvae fedon the samehost species (Table 1).TheBMG nymphs which fed on R. norvegicus or on C.callosus required longer premolt periods (P , 0.05)than didCSPnymphs reared on the samehost species.

Figures 1 and 2 show the larval detachment rhythmand the dynamics of larva-to-nymph molt. Using R.norvegicus as hosts for both tick colonies, the Þrstnymph appeared 3 or 4 d after the last engorged larvaehad detached and 100% of the nymphs emerged '25d after larval infestation. In contrast, using C. callosusas hosts for larvae, the Þrst nymph appeared whilelarval ticks were still feeding. BMG and CSP nymphsÞrst appeared when 98 and 74% of the total engorgedlarvaehaddetached fromC. callosus, respectively, and100% of the nymphs emerged '25 d after larval in-festation. Figs. 3 and 4 show the nymphal detachmentrhythm and the dynamics of the nymph-to-adult molt.For both tick colonies, using R. norvegicus or C. cal-losus as hosts, the Þrst adult appeared after all nymphshad detached, and 100% of the adults emerged '30 dafter the nymphal infestation.

Diurnal Detachment. With the exception of theimmature CSP ticks fed on R. novergicus, whichshowed similar (P . 0.05) detachment during day and

Table 1. Biological data of immature stages I.(I.) loricatus from two localities in Brazil

Sitea Stage Hostb No. exposedNo.

recovered(%)

Feeding period, d % Diurnaldetachment

No. molted(%)

Premolt period, d

Mean 6 SD Range Mean 6 SD Range

BMG Larvae Rn 3,000 702 (23.4)a 6.43 6 0.85a 5Ð9 63.7a 481 (16.0)a 10.24 6 1.68a 7Ð22Cc 3,000 959 (32.0)b 8.03 6 1.99b 5Ð16 89.1b 836 (27.9)b 8.99 6 1.37b 8Ð20

Nymphs Rn 410 33 (8.0)A 6.33 6 0.78A 5Ð9 66.7A,C 17 (4.1)A 22.35 6 1.22A 21Ð26Cc 410 47 (11.5)B 6.26 6 1.09A 5Ð9 89.4B 34 (8.0)B 21.38 6 1.52A,B 19Ð25

CSP Larvae Rn 2,875 360 (12.5)c 6.82 6 0.93c 5Ð9 46.7c 237 (8.2)c 10.64 6 2.23c 8Ð19Cc 2,875 422 (14.7)d 11.13 6 2.58d 5Ð19 64.4a 289 (10.0)d 10.15 6 7.00d 8Ð19

Nymphs Rn 255 71 (27.8)C 7.49 6 1.04B 5Ð10 54.0A 58 (22.7)C 20.60 6 1.60B 18Ð26Cc 335 66 (19.7)C 8.26 6 1.52B 5Ð13 68.0C 65 (19.4)C 18.78 6 1.24C 16Ð21

Means in the same column followed by different letters are signiÞcantly different (P , 0.05). Larvae, lower-case letters; nymphs, capitalletters.

a BMG, Belo Horizonte County, state of Minas Gerais; CSP, Cotia County, state of Sao Paulo.b Rn, Rattus norvegicus; Cc, Calomys callosus.

Fig. 1. Dropping-off andmolting rhythmof engorged I. loricatus larvae fromBeloHorizonteCounty, state ofMinasGerais(BMG). Triangles and circles represent larvae fed on Rattus norvegicus and Calomys callosus, respectively.

716 JOURNAL OF MEDICAL ENTOMOLOGY Vol. 37, no. 5

night, the majority of larvae and nymphs detachedfrom the rodents during the day (Table 1). Within thesame colony, the percentage detachment of ticks fromC. callosusduring thedaywas signiÞcantlyhigher (P,0.05) than that observed for ticks fed on R. norvegicus.

Recovered and Molted Tick Percentages. For ticksof both BMG and CSP colonies, the percentage ofrecovered or molted ticks in relation to the initialnumber of unfed ticks released on the hosts was sig-niÞcantly higher (P , 0.05) for larvae and nymphs fedon C. callosus than those recorded for R. norvegicus,except for CSP nymphs, for which a greater percent-age was recovered or molted when fed on R. rattus(not signiÞcant, P . 0.05) (Table 1). However, thepercentageof recovered ticks that successfullymoltedafter feeding on C. callosus was, in all cases, signiÞ-cantly higher (P , 0.05) than that recorded for im-mature ticks fed on R. norvegicus. For BMG ticks fedon C. callosus and R. novergicus, 87.2 and 68.5% of therecovered larvae, respectively, successfully molted tonymphs, and 68.5 and 65.8% of the recovered CSPlarvae that fed on C. callosus and R. novergicus, re-spectively, molted to nymphs. Seventy-two and 51.5%

of BMG nymphs that fed on C. callosus and R. nover-gicus, respectively, successfully molted to adults, and98.5 and 81.7% of CSP nymphs successfully moltedafter feeding on C. callosus and R. novergicus, respec-tively.

First and Second Tick Generations. The individualbiological data for the Þrst and second generation ofimmature ticks from the BMG or CSP colonies (datanot shown) were similar to those shown in Table 1.

Adult Stage. As a result of the two tick generationsobtained in the laboratory, 51 BMG adult ticks (25males, 26 females) and 65 CSP adult ticks (31 males,34 females) emerged. Once the adult ticks were sub-mitted to mating conditions, all adult ticks were al-lowed to feed on one of the opossums. Fourteen of 16BMG females that were recovered oviposited, andfrom the CSP tick colony, seven of 17 females pro-duced eggs. The weight of one egg was 0.008 mg 60.005 forBMGticks and0.006mg60.042 forCSP ticks.One male tick from the BMG colony and two malesfrom CSP were recovered.

Table 2 contains the biological data of females andeggs that were obtained from the two generations of

Fig. 3. Dropping-off and molting rhythm of engorged I. loricatus nymphs from Belo Horizonte County, state of MinasGerais (BMG). Triangles and circles represent nymphs fed on Rattus norvegicus and Calomys callosus, respectively.

Fig. 2. Dropping-off and molting rhythm of engorged I. loricatus larvae from Cotia County, state of Sao Paulo (CSP).Triangles and circles represent larvae fed on Rattus norvegicus and Calomys callosus, respectively.

September 2000 SCHUMAKER ET AL.: LIFE CYCLE OF I. (I.) loricatus 717

the BMG and CSP tick colonies. CSP females fedfaster, but their oviposition period was longer com-pared with the BMG females (P , 0.05).

Table 3 shows the biological data obtained for fe-males and eggs from the Þrst and second generationsof the BMG tick colony. Females from the second tickgeneration (n 5 5) had signiÞcantly (P , 0.05) lowermean numbers of eggs per female, egg hatching per-centage, and egg production efÞciency indexes com-pared with those females from the Þrst generation(n 5 9). Five females from the Þrst generation of theCSP ticks and two females from the second generationlaidviableeggs; becauseof this lownumberof females,no comparative test was conducted.

Life Cycle. The total duration of the life cycle wasestimated by adding the duration of larval andnymphal stages (Table 1) and the duration of feeding,preovipostion, and egg incubation periods (Table 2)plus themean prefeeding periods of 50, 30 and 40 d forlarvae, nymphs, and adults, respectively. For ticks inthe BMG colony, the mean duration of the life cycleswas 222 d, using either R. norvegicus or C. callosus ashosts for immature ticks, respectively. The ticks in theCSP colony had a mean of 218 or 221 d after feedingon R. norvegicus or C. callosus, respectively. The over-all sex ratio was 1:1 for the BMG colony.

Discussion

Aragao (1936) was unsuccessful in rearing I. (I.)loricatus larvae on opossum hosts but he did obtain afew engorged larvae, some of which molted to thenymphal stage, using white rats as hosts. In earlierwork,we failed to get the adults to feed on rats and thelarvae on opossums (M.B.L., unpublished data). Thus,we used rodents and opossums to feed immature andadult ticks, respectively. This provided adequate bio-logical data fromwhich the life cycle of I. (I.) loricatuscould be estimated under laboratory conditions.

Because in Brazil several Sigmodontinae rodentspecies have been found naturally infested by I. (I.)loricatus larvae and nymphs (Barros-Battesti 1998,Abel 1999), we used C. callosus, a native sigmodontinefrom South America (Musser and Carleton 1993), asan experimental wild host for immature ticks.

Despite the signiÞcantdifferences in thedurationoffeeding or premolt periods between immature ticksfed on R. norvegicus or C. callosus (Table 1), the larvalfeeding period (including the prefeeding period) wasestimated to be '2 mo. The nymphal feeding periodwas '2 mo, independent of the origin of the colony(BMG or CSP) or the host used. In the majority of theobservations (Table 1), the recovered and molted

Fig. 4. Dropping-off and molting rhythm of engorged of I. loricatus nymphs from Cotia county, state of Sao Paulo (CSP).Triangles and circles represent nymphs fed on Rattus norvergivus and Calomys callosus, respectively.

Table 2. Biological data of I.(I.) loricatus females from two localities in Brazil

FactorBMG (n 5 14) CSP (n 5 7)

Mean 6 SD Range Mean 6 SD Range

Feeding period, d 11.43 6 2.85 9Ð14 8.71 6 1.79* 6Ð12Weight of engorged female, mg 327.43 6 48.09 228Ð409 313.14 6 54.19 256Ð389Preoviposition period, d 6.29 6 1.14 4Ð8 5.57 6 0.53 5Ð7Oviposition period, d 31.36 6 5.65 21Ð42 38.13 6 8.59* 33Ð48Weight of deposited eggs, mg 135.00 6 46.96 50Ð214 138.43 6 35.45 84Ð177Number of eggs/female 1,623.14 6 709.14 625Ð2,675 2,232.57 6 571.85 1,355Ð2,855Egg incubation period, d 38.79 6 2.83 34Ð42 38.43 6 2.64 35Ð42Hatching time, d 23.43 6 8.71 9Ð34 20.14 6 7.69 11Ð35% egg hatch 56.79 6 25.39 5Ð90 42.86 6 27.06 10Ð80Egg production efÞciency 40.79 6 11.62 23.58Ð54.85 44.05 6 8.35 32.18Ð53.43

* SigniÞcantly different from BMG females (P , 0.05).

718 JOURNAL OF MEDICAL ENTOMOLOGY Vol. 37, no. 5

percentageswere signiÞcantlyhigher (P,0.05)whenlarvae or nymphs were fed on C. callosus. Our dataindicated thatC. callosuswas amore suitable host thanthe laboratory host.

In spite of our daily examinations of the cage, thewood shavings, and the host animal food, only 5% and55% of the total males and females, respectively, werefoundafterbeingplacedon theopossums. It is possiblethat the ticks were eaten by the host as observed byAubert (1981),who reported that engorged females ofI. (Pholeoixodes) rugicollis were often eaten by thehost ferret (Mustela furo), and in some cases 40Ð60%of the exposed females were not recovered.

Although the CSP female ticks fed faster and re-quired longer oviposition periods than the BMG fe-males, other reproductive parameters were similar(Table 2). The total number of eggs that were depos-ited per female ranged from 625 to 2,675 and from1,355 to 2,855 for I. (I.) loricatus BMG and CSP ticks,respectively. These values were similar to the eggnumbers reported by Aragao (1936) for this tick spe-cies (1,300 eggs per female), and were similar to thosereported for other species of the subgenus Ixodes(Arthur and Snow 1968, Oliver et al. 1987, Wilson etal. 1990, Banks et al. 1998). The average I. (I.) loricatusegg production efÞciency ('41Ð44%) was close tothat observed for mated females of I. (P.) hexagonous(40.26%), which tended to increase egg numbers rel-ative to the engorgement weight (Toutoungi et al.1995).

All I. (I.) loricatus females from both the BMG andCSP tick colonies were exposed to the mating condi-tions before infestation, and some male ticks could beseen wandering or attached to the skin of opossumsduring the adult feeding period. However, 12 of 33partially or fully engorged females (weight '269 mg,range 204Ð388 mg) either failed to lay eggs or ovipos-ited only small batches of nonviable eggs (mass eggweight '49 mg, range 29Ð78 mg). These unsuccessfulfemales were probably unfertilized as observed invirgin females of I. ruggicolis and I. hexagonous, whichcan engorge and lay a few viable eggs (Aubert 1981,Toutoungi et al. 1995).

Most Ixodes biology studies have used numerousticks collected from the Þeld to start a laboratorycolony (Farkas and Surgeoner 1991, Toutoungi et al.

1993, Banks et al. 1998). We used only a single en-gorged female to start each colony (BMGorCSP) andno other ticks were collected. The inbreeding of thesecond generation of the F1 progeny may explain thedrastic reduction in fertility observed on the BMGfemales. In addition,weusedÞeld-collected opossumsto feed the adult ticks and their previous contact withticks was not known. Host immune response to tickbites may lead to a state of resistance against ticks,resulting innegative reproduction indexes (Bowessid-jaou et al. 1977, Girardin and Brossard 1990); this mayhave adversely affected the results obtained in thisstudy.

Thisworkprovides life cycle informationof two tickcolonies initiated from two geographically isolatedfemales. The Þeld-collected BMG and CSP femaleswere identiÞed as I. (I.) loricatus using the taxonomickey published by Fairchild et al. (1966). Had we hadused the Brazilian key published by Aragao and Fon-seca (1961), the Þeld-collected BMG female wouldhave been identiÞed as Ixodes (Ixodes) didelphidisFonseca andAragao.Bymorphological examinationofthe shapeof the spiracular plates of theF1 andF2 adultticks of BMGandCSP colonies,we observed that bothcolonies yielded adult females that could be identiÞedas I. (I.) loricatus but others as I. (I.) didelphidis.However, all adultmales fromboth tick colonies couldbe identiÞed as I. (I.) didelphidis according to thedescription provided by Fonseca and Aragao (1952).Fairchild et al. (1966) considered that the originaldescription of I. didelphidis by Fonseca and Aragao(1952) probably referred to a local population, incontrast with I. loricatus ticks that had been reportedfrom Mexico and southward to Argentina. As long asthe validation of the species I. (I.) didelphidis is un-resolved, we consider the two populations studied asa single species.

Acknowledgments

We thank J. Klotzel (Instituto deMedicinaTropical/USP)for furnishing Calomys callosus, and S. H. Lima and P.C.F.Silva for technical assistance. We also thank the InstitutoBrasileiro do Meio Ambiente e dos Recursos Naturais Reno-vaveis (IBAMA) for permitting the capture, handling, andrelease of wild animals. This work was supported by the

Table 3. Biological data of I.(I.) loricatus females from Belo Horizonte, state of Minas Gerais, Brazil

FactorGeneration 1 (n 5 9) Generation 2 (n 5 5)

Mean 6 SD Range Mean 6 SD Range

Feeding period, d 12.30 6 2.87 9Ð14 9.25 6 1.26 9Ð14Weight of engorged female, mg 324.67 6 50.81 228Ð392 332.40 6 48.02 280Ð409Preoviposition period, d 6.20 6 1.32 4Ð8 6.50 6 0.57 5Ð7Oviposition period, d 32.80 6 5.20 27Ð42 27.75 6 5.74 21Ð35Weight of deposited eggs, mg 152.10 6 36.86 99Ð214 104.20 6 51.03 50Ð172Number of eggs/female 1,901.33 6 460.95 1,300Ð2,675 1,122.40 6 849.61* 625Ð2,150Egg incubation period, d 37.90 6 2.28 34Ð41 41.00 6 3.16 15Ð26Hatching time, d 24.80 6 9.74 9Ð34 20.00 6 4.69 15Ð26% egg hatch 67.22 6 20.33 30Ð90 38.00 6 24.14* 5Ð60Egg production efÞciency 46.58 6 6.83 36.17Ð52.72 30.36 6 11.58* 17.85Ð43.33

* SigniÞcantly different from the Þrst generation (P , 0.05).

September 2000 SCHUMAKER ET AL.: LIFE CYCLE OF I. (I.) loricatus 719

Brazilian research agencies Conselho Nacional de Desen-volvimento CientõÞco e Tecnologico (CNPq) and Fundacaode Amparo a Pesquisa do Estado de Sao Paulo (FAPESP).

References Cited

Abel, I. S. 1999. Investigacoes preliminares sobre espiro-quetõdeos e seus potenciais vetores procedentes dareserva ßorestal do Morro Grande, Municõpio de CotiaÐSP. Ph.D. dissertation, ICB-Universidade de Sao Paulo.

Aragao, H. B. 1936. Ixodidas brasileiros e de alguns paizeslimitrophes. Mem. Inst. Oswaldo Cruz. 31: 759Ð843.

Aragao, H. B., and F. Fonseca. 1961. Notas de ixodologia.VIII. Lista e chave para os representantes da fauna ixod-ologica brasileira. Mem. Inst. Oswaldo Cruz. 59: 115Ð129.

Arthur, D. R., and K. R. Snow. 1968. Ixodes pacificus Cooleyand Kohls, 1943: its life-history and occurrence. Parasi-tology 58: 893Ð906.

Aubert, M.F.A. 1981. Breeding of the tick Ixodes (Pholeoix-odes) rugicollis (Acari:Ixodidae) under laboratory con-ditions. J. Med. Entomol. 18: 324Ð327.

Banks, C. W., J. H. Oliver, Jr., J. B. Philips, and K. L. Clark.1998. Life cycle of Ixodes minor (Acari: Ixodidae) in thelaboratory. J. Med. Entomol. 35: 496Ð499.

Barros-Battesti, D. M. 1998. Estudos de carrapatos e peque-nos mamõferos silvestres naturalmente infectados comespiroquetas semelhantes a borrelia, no municõpio deItapevi, Estado de Sao Paulo. M.S. thesis, FSP-Univer-sidade de Sao Paulo.

Bennett, G. F. 1974. Oviposition of Boophilus microplus(Canestrini) (Acarina: Ixodidae). Acarologia 16: 1652Ð1661.

Bowessidjaou, J., M. Brossard, and A. Aeschlimann. 1977.Effects and duration of resistance acquired by rabbits onfeeding and egg laying in Ixodes ricinus L. Experientia 33:548Ð550.

Clifford, C. M., D. E. Sonenshine, J. E. Keirans, and G. M.Kohls. 1973. Systematics of the subfamily Ixodinae(Acarina: Ixodidae). 1. The subgenera of Ixodes. Ann.Entomol. Soc. Am. 66: 489Ð500.

Cooley, R. A., and G. M. Kohls. 1945. The genus Ixodes inNorth American Bulletin 184, National Institutes ofHealth, Rocky Mountain Laboratory, Hamilton, MT.

Drummond, R. O., S. E. Ernst, J. L. Trevino, W. J. Gladney,and O. H. Graham. 1973. Boophilus annulatus and B.microplus: laboratory tests of insecticides. J. Econ. Ento-mol. 66: 130Ð133.

Fairchild, G. B., G. M. Kohls, and V. Tipton. 1966. The ticksof Panama, pp. 167Ð219. In R. L. Wenzel and V. J. Tipton[eds], Ectoparasites of Panama. FieldMuseumofNaturalHistory, Chicago, IL.

Farkas, M. J., and G. A. Surgeoner. 1991. Developmentaltimes and fecundity of Ixodes cookei Packard (Acari:Ixodidae) under laboratory conditions. Can. Entomol.123: 1Ð12.

Fonseca, F., and H. B. Aragao. 1952. Notas de Ixodologia. II.Uma nova especie do genero Amblyomma e uma novaespecie do genero Ixodes (Acari: Ixodidae). Mem. Inst.Oswaldo Cruz. 50: 713Ð726.

Girardin, P., and M. Brossard. 1990. Rabbits infested withIxodes ricinus L. adults: effects of a treatment with cy-closporin A on the biology of ticks fed on naive andimmune hosts. Ann. Parasitol. Hum. Comp. 65: 262Ð266.

Gray, J. S. 1998. The ecology of ticks transmitting Lymeborreliosis. Exp. Appl. Acarol. 22: 249Ð258.

Musser, G. G., and M. D. Carleton. 1993. Order Rodentia,pp: 501Ð806. In D. E. Wilson and D. M. Reeder [eds.],Mammal species of theworld: a taxonomic and geograph-ical reference. Smithsonian Institution, Washington, DC.

Nuttal, G.H.F., and C. Warburton. 1911. TicksÑa mono-graph of the Ixodoidea. Part II. The genus Ixodes. Cam-bridge University Press, London.

Oliver, J. H., Jr. 1974. Symposium on reproduction of ar-thropods of medical and veterinary importance. IV. Re-production in ticks (Ixodoidea). J. Med. Entomol. 11:26Ð34.

Oliver, J. H., Jr. 1999. Lyme borreliosis in the southernUnited States: a review. J. Parasitol. 82: 926Ð935.

Oliver, J. H., Jr., J. E. Keirans, D. R. Lavender, and H. J.Hutcheson. 1987. Ixodes afinnis Newmann (Acari: Ixo-didae): new host and distribution records, description ofimmatures, seasonal activities in Georgia, and laboratoryrearing. J. Parasitol. 73: 646Ð652.

Toutoungi, L. N., L. Gern, and A. Aeschlimann. 1993. Biol-ogy of Ixodes (Pholeoixodes) hexagonus under laboratoryconditions. Part I. Immature stages. Exp. Appl. Acarol. 17:655Ð662.

Toutoungi, L. N., L. Gern, and A. Aeschlimann. 1995. Biol-ogy of Ixodes (Pholeoixodes) hexagonus under laboratoryconditions. Part II. Effect of mating on feeding and fe-cundity of females. Exp. Appl. Acarol. 19: 233Ð245.

Wilson,M.L., T. S. Litwin, T.A.Gavin,M.C.Capkanis,D.C.Maclean, and A. Spielman. 1990. Host-dependent dif-ferences in feeding and reproduction of Ixodes dammini(Acari: Ixodidae). J. Med. Entomol. 27: 945Ð954.

Yoshinari, N. H., A. C. Steere, P.J.L. Barros, F.C.M. Cruz, M.Mendonca, L. K. Oyafuso, L. Levy, and W. Cossermelli.1993a. Lyme disease in Brazil: report of Þve cases. Rev.Esp. Reumat. 20(suppl.): 156.

Yoshinari, N. H., P.J.L. Barros, R. Stelling, D. Baggio, P.Yassuda, and W. Cossermelli. 1993b. Epidemiologicalstudy of Lyme disease in Cotia. Rev. Esp. Reumatol.20(suppl.): 474.

Yoshinari, N. H., V.L.N. Bonoldi, D. M. Barros-Battesti, andT.T.S. Schumaker. 1999. Doenca de Lyme-sõmile noBrasil. Rev. Bras. Reumatol. 29: 1Ð2.

Received for publication21July 1999; accepted12May2000.

720 JOURNAL OF MEDICAL ENTOMOLOGY Vol. 37, no. 5