Laboratory Studies on the Life Cycle of Amblyomma marmoreum (Acari: Ixodidae) on Two Different Hosts LAURA JANE FIELDEN,* SOLOMON MAGANO, AND YIGAL RECHAV Department of Biology, Medical University of Southern Africa, P.O. Medunsa 0204, Republic of South Africa
KEY WORDS Arachnida, Amblyomma marmoreum, life cycle, hosts
Amblyomma marmoreum Koch, 1844 occurs over much of southeastern Africa and is particularly common in the Eastern Cape province of South Africa (Theiler & Salisbury 1959). Adults are specific for reptiles, particularly tortoises (Norval 1975, Petney & Horak 1988, Ghirotti & Mwanaumo 1989, Walker 1991), whereas immature stages utilize a wide variety of reptiles, birds, mammals, and domestic stock as hosts (Horak & Knight 1986, Horak & Williams 1986, Horak et al. 1987, Petney et al. 1987). Amblyomma marmoreum has recently been found to be a suitable laboratory vector of the rickettsia Cowdria ruminantium, the cause of heartwater in domestic ruminants in South Africa (Bezuidenhout 1987). However, the natural role of A. marmoreum in the transmission of heartwater remains unclear, although Petney & Horak (1988) have suggested that this tick may be implicated in direct transmission of C. ruminantium to domestic stock via the immature stages. In light of the potential economic importance of A. marmoreum, any study on the biology of this species is of significance. Some information on the ecology of A. marmoreum and its life cycle under laboratory conditions is available (Norval 1975, Rechav 1982, Rechav et al. 1987). However, certain details of the life cycle, including the development of adult ticks on mammal hosts, have not yet been documented. 1 Current address: Department of Physiology, University of the North, Private Bag XI106, Sovenga 0727, Republic of South Africa.
Considering the host specificity of adult ticks in comparison with immature stages, a study of the effect of various hosts on the life cycle of this tick species seems appropriate. This work provides details on the life cycle of A. marmoreum under controlled laboratory conditions on reptilian (tortoise) and mammalian (guinea pig) hosts. Materials and Methods Ticks used were either from laboratory stocks or collected from leopard tortoises (Geochelone pardalis) at the National Zoological Gardens, Pretoria, South Africa. Larvae, nymphs, and adults were fed on female tortoises and on 3-moold naive female Dunkin-Hartley guinea pigs maintained individually in the laboratory (25°C, natural light regime, ambient humidity). Larvae, nymphs, and adults fed on guinea pigs were released into cotton pockets glued onto the shaved skin of the host. Attachment of adults was induced by smearing the body contents of a partially fed adult male A. marmoreum onto the back of the host. Ticks fed on tortoises were released directly onto the head and forelimbs of the host. In totals, 100 larvae and 50 nymphs were pulled daily from three guinea pigs (equal numbers taken from each host), and totals of 100 larvae and 15 nymphs were removed at 3-d intervals from four tortoises. Pulling of larvae and nymphs from hosts continued until the start of the drop-off period. Ticks were weighed on a Sartorius 1801 electric balance immediately after their removal.
0022-2585/92/0750-0756$02.00/0 © 1992 Entomological Society of America
Downloaded from http://jme.oxfordjournals.org/ at University of Nebraska-Lincoln Libraries on June 8, 2016
J. Med. Entomol. 29(5): 750-756 (1992) ABSTRACT A comparative study was made of the life cycle of the tortoise tick, Amblyomma marmoreum Koch, on tortoises and guinea pigs under laboratory conditions. At 25°C and 85% RH with natural day length, duration of off-host stages (preoviposition, oviposition, incubation, and premolt) was similar for ticks fed on both hosts. Delay in preoviposition (up to 90 d) was observed in some gravid females. Larvae, nymphs, and adults had longer feeding periods on tortoises than on guinea pigs. Adult females fed on tortoises had greater engorgement weight and ovipositional capacity than ticks fed on guinea pigs. It is suggested that the shortened feeding period of immature stages on mammalian hosts together with the occurrence of morphogenic diapause may be more important than previously recognized in determining whether the life cycle of A. marmoreum is completed in 1 or 2 yr under natural conditions.
September 1992
FIELDEN ET AL.: L I F E CYCLE
Table 1. Preoviposition period of A. marmoreum collected from tortoises at various periods front October 1990 to January 1991 Month
n
Preoviposition, d Mean ± SD Range
October November December January
23 23 9 12
21.7:t 7.4 22.4:t 8.0 55.4:t27.6 22.7:t 5.6
14-41 12-48 20-95 14-32
Results Preoviposition. The preoviposition period of 67 females collected from free-living and laboratory-maintained tortoises was extremely variable, ranging from 13 to 95 d. Engorged females that dropped during December had significantly longer preoviposition periods (ANOVA; F = 20.01, P < 0.001) compared with females that
marmoreum
751
dropped during October, November, and January (Table 1). The preoviposition period for five females collected from guinea pigs during a 10-d period from the end of November to early December ranged from 33 to 93 d (71.0 ± 27.6 d). Low yield of engorged females from guinea pigs for measurement of the preoviposition period resulted primarily from failure of ticks to attach to the mammalian host. Only 8 of 15 females successfully completed their blood meal on guinea pigs; three of these eight females subsequently died before oviposition. Oviposition. The average daily oviposition rate of 10 females fed on tortoises is shown in Fig. 1. Oviposition reached a peak on day 8, then slowly declined with 75% of the eggs laid within the first 18 d. The mean oviposition period was 31.1 ± 8.7 d compared with 29.8 ± 2.4 d for five females fed on guinea pigs. The average number of eggs produced by females from tortoise hosts was 11,720.0 ± 5,547.5 (n = 20) compared with 4,552.8 ± 1,728.0 (n = 5) from guinea pig hosts. A highly significant positive correlation (r = 0.91, P < 0.001) was found between the weight of eggs laid and the weight of the engorged females (Fig. 2). From the x axis intercept of the regression fitted to the data, it was estimated that 0.871 g of the weight of the females was utilized for metabolic purposes other than egg production.
9OO
BOO-
§ CO
7OO
+
6OO-
en 5OO400 ••
3OO • 2 200 • 1OO-
I 5
1O
15
20
25
3O
1
35 A-O
Days Fig. 1. Daily oviposition of A. marmoreum collected from tortoises. Vertical lines represent SDs of the mean (based on 10 females).
Downloaded from http://jme.oxfordjournals.org/ at University of Nebraska-Lincoln Libraries on June 8, 2016
All engorged stages that dropped from the host were weighed, placed in glass desiccators above a saturated solution of KC1 to provide a relative humidity of 85 ± 5% (Winston & Bates 1960), and maintained at 25°C with a natural light cycle. Daily checks were made to record the number of eggs laid and the time taken by larvae, nymphs, and adults to molt. Data are presented as mean ± SD.
OF Amblyomma
752
Vol. 29, no. 5
JOURNAL OF MEDICAL ENTOMOLOGY 3.O
Y = 0.6365x - 0.4506 r = 0.91 2.5
•
2
1.5
• •
• .
1
0.5
• 1
. •
2
3
4
5
Weight of females (g)
Fig. 2. Relationship between weight of eggs and weight of engorged females of A. marmoreum collected from tortoise and guinea pig hosts.
Incubation Period. The incubation period (defined as the time from beginning of oviposition to the emergence of the first larva) was 62.0 ± 2.7 d for 30 ticks fed on tortoises and 57.7 ± 6.0 d for three females collected from guinea pigs. Feeding Period and Weight of Larvae. Larvae required a feeding period of 16.8 ± 3.2 d (n = 836) on tortoises and 8.8 ± 1.7 d on guinea pigs (n = 432). Daily drop-off of engorged larvae reached peaks on days 7 and 15 from guinea pigs and tortoises, respectively (Fig. 3). The feeding pattern was similar for larvae on both hosts: a phase of slow gain in weight (days 1—4 on guinea pigs, days 1-9 on tortoises) was followed by a phase of rapid engorgement and weight gain before dropping off the host (Fig. 4). The mean drop-off weight of larvae fed on guinea pigs was 2.64 ± 0.22 mg compared with 2.55 ± 0.22 mg for larvae fed on tortoises. The molting period of the engorged larvae to nymphs fed on guinea pigs was 25.9 ± 3.0 d (n = 239) and that of larvae fed on tortoises was 22.1 ± 2.8 d (n = 340).
A 6 8 10 12 2 Days after attachment Fig. 4. Changes in weight (mean ± SD) of larvae of A. marmoreum during their feeding period (based on samples of 10 larvae each). —, tortoise; —, guinea pig.
Feeding Period and Weight of Nymphs. The average feeding period of nymphs was 55.0 ± 15.3 d (n = 76) on tortoises and 7.7 ± 4.3 d (n = 195) on guinea pigs. Peak drop-off from guinea pigs occurred 8 d after attachment, then rapidly declined (Fig. 5). Drop-off from tortoises started 18 d after attachment and continued until day 77 (Fig. 5). No peak drop-off period was evident. The pattern of feeding for nymphs on guinea pigs (Fig. 6) was similar to that of larvae with a period of slight gain in weight (days 1-5), followed by a phase of rapid weight gain which coincided with the start of drop-off of replete nymphs. Feeding on tortoises was also characterized by a period of slow weight gain (Fig. 6). However, the phase of rapid weight gain before repletion could not be detected because engorged nymphs dropped in low numbers (usual8O 7O •
I y/6
8 1O 12 14- 1 6 18 2O 22 24- 26 Days after attachment
Fig. 3. Daily drop-off of A. marmoreum larvae. LTD, tortoise; WM, guinea pig.
6
„n
8 1O 15 25 35 4-5 55 65 75 Days after attachment
Fig. 5. Daily drop-off of A. marmoreum nymphs. LTD, tortoise; • • , guinea pig.
Downloaded from http://jme.oxfordjournals.org/ at University of Nebraska-Lincoln Libraries on June 8, 2016
0
•
.
September 1992
FIELDEN E T A L . : L I F E CYCLE
OF Amblyomma
marmoreum
753
1
KEY WORDS Arachnida, Amblyomma marmoreum, life cycle, hosts
Amblyomma marmoreum Koch, 1844 occurs over much of southeastern Africa and is particularly common in the Eastern Cape province of South Africa (Theiler & Salisbury 1959). Adults are specific for reptiles, particularly tortoises (Norval 1975, Petney & Horak 1988, Ghirotti & Mwanaumo 1989, Walker 1991), whereas immature stages utilize a wide variety of reptiles, birds, mammals, and domestic stock as hosts (Horak & Knight 1986, Horak & Williams 1986, Horak et al. 1987, Petney et al. 1987). Amblyomma marmoreum has recently been found to be a suitable laboratory vector of the rickettsia Cowdria ruminantium, the cause of heartwater in domestic ruminants in South Africa (Bezuidenhout 1987). However, the natural role of A. marmoreum in the transmission of heartwater remains unclear, although Petney & Horak (1988) have suggested that this tick may be implicated in direct transmission of C. ruminantium to domestic stock via the immature stages. In light of the potential economic importance of A. marmoreum, any study on the biology of this species is of significance. Some information on the ecology of A. marmoreum and its life cycle under laboratory conditions is available (Norval 1975, Rechav 1982, Rechav et al. 1987). However, certain details of the life cycle, including the development of adult ticks on mammal hosts, have not yet been documented. 1 Current address: Department of Physiology, University of the North, Private Bag XI106, Sovenga 0727, Republic of South Africa.
Considering the host specificity of adult ticks in comparison with immature stages, a study of the effect of various hosts on the life cycle of this tick species seems appropriate. This work provides details on the life cycle of A. marmoreum under controlled laboratory conditions on reptilian (tortoise) and mammalian (guinea pig) hosts. Materials and Methods Ticks used were either from laboratory stocks or collected from leopard tortoises (Geochelone pardalis) at the National Zoological Gardens, Pretoria, South Africa. Larvae, nymphs, and adults were fed on female tortoises and on 3-moold naive female Dunkin-Hartley guinea pigs maintained individually in the laboratory (25°C, natural light regime, ambient humidity). Larvae, nymphs, and adults fed on guinea pigs were released into cotton pockets glued onto the shaved skin of the host. Attachment of adults was induced by smearing the body contents of a partially fed adult male A. marmoreum onto the back of the host. Ticks fed on tortoises were released directly onto the head and forelimbs of the host. In totals, 100 larvae and 50 nymphs were pulled daily from three guinea pigs (equal numbers taken from each host), and totals of 100 larvae and 15 nymphs were removed at 3-d intervals from four tortoises. Pulling of larvae and nymphs from hosts continued until the start of the drop-off period. Ticks were weighed on a Sartorius 1801 electric balance immediately after their removal.
0022-2585/92/0750-0756$02.00/0 © 1992 Entomological Society of America
Downloaded from http://jme.oxfordjournals.org/ at University of Nebraska-Lincoln Libraries on June 8, 2016
J. Med. Entomol. 29(5): 750-756 (1992) ABSTRACT A comparative study was made of the life cycle of the tortoise tick, Amblyomma marmoreum Koch, on tortoises and guinea pigs under laboratory conditions. At 25°C and 85% RH with natural day length, duration of off-host stages (preoviposition, oviposition, incubation, and premolt) was similar for ticks fed on both hosts. Delay in preoviposition (up to 90 d) was observed in some gravid females. Larvae, nymphs, and adults had longer feeding periods on tortoises than on guinea pigs. Adult females fed on tortoises had greater engorgement weight and ovipositional capacity than ticks fed on guinea pigs. It is suggested that the shortened feeding period of immature stages on mammalian hosts together with the occurrence of morphogenic diapause may be more important than previously recognized in determining whether the life cycle of A. marmoreum is completed in 1 or 2 yr under natural conditions.
September 1992
FIELDEN ET AL.: L I F E CYCLE
Table 1. Preoviposition period of A. marmoreum collected from tortoises at various periods front October 1990 to January 1991 Month
n
Preoviposition, d Mean ± SD Range
October November December January
23 23 9 12
21.7:t 7.4 22.4:t 8.0 55.4:t27.6 22.7:t 5.6
14-41 12-48 20-95 14-32
Results Preoviposition. The preoviposition period of 67 females collected from free-living and laboratory-maintained tortoises was extremely variable, ranging from 13 to 95 d. Engorged females that dropped during December had significantly longer preoviposition periods (ANOVA; F = 20.01, P < 0.001) compared with females that
marmoreum
751
dropped during October, November, and January (Table 1). The preoviposition period for five females collected from guinea pigs during a 10-d period from the end of November to early December ranged from 33 to 93 d (71.0 ± 27.6 d). Low yield of engorged females from guinea pigs for measurement of the preoviposition period resulted primarily from failure of ticks to attach to the mammalian host. Only 8 of 15 females successfully completed their blood meal on guinea pigs; three of these eight females subsequently died before oviposition. Oviposition. The average daily oviposition rate of 10 females fed on tortoises is shown in Fig. 1. Oviposition reached a peak on day 8, then slowly declined with 75% of the eggs laid within the first 18 d. The mean oviposition period was 31.1 ± 8.7 d compared with 29.8 ± 2.4 d for five females fed on guinea pigs. The average number of eggs produced by females from tortoise hosts was 11,720.0 ± 5,547.5 (n = 20) compared with 4,552.8 ± 1,728.0 (n = 5) from guinea pig hosts. A highly significant positive correlation (r = 0.91, P < 0.001) was found between the weight of eggs laid and the weight of the engorged females (Fig. 2). From the x axis intercept of the regression fitted to the data, it was estimated that 0.871 g of the weight of the females was utilized for metabolic purposes other than egg production.
9OO
BOO-
§ CO
7OO
+
6OO-
en 5OO400 ••
3OO • 2 200 • 1OO-
I 5
1O
15
20
25
3O
1
35 A-O
Days Fig. 1. Daily oviposition of A. marmoreum collected from tortoises. Vertical lines represent SDs of the mean (based on 10 females).
Downloaded from http://jme.oxfordjournals.org/ at University of Nebraska-Lincoln Libraries on June 8, 2016
All engorged stages that dropped from the host were weighed, placed in glass desiccators above a saturated solution of KC1 to provide a relative humidity of 85 ± 5% (Winston & Bates 1960), and maintained at 25°C with a natural light cycle. Daily checks were made to record the number of eggs laid and the time taken by larvae, nymphs, and adults to molt. Data are presented as mean ± SD.
OF Amblyomma
752
Vol. 29, no. 5
JOURNAL OF MEDICAL ENTOMOLOGY 3.O
Y = 0.6365x - 0.4506 r = 0.91 2.5
•
2
1.5
• •
• .
1
0.5
• 1
. •
2
3
4
5
Weight of females (g)
Fig. 2. Relationship between weight of eggs and weight of engorged females of A. marmoreum collected from tortoise and guinea pig hosts.
Incubation Period. The incubation period (defined as the time from beginning of oviposition to the emergence of the first larva) was 62.0 ± 2.7 d for 30 ticks fed on tortoises and 57.7 ± 6.0 d for three females collected from guinea pigs. Feeding Period and Weight of Larvae. Larvae required a feeding period of 16.8 ± 3.2 d (n = 836) on tortoises and 8.8 ± 1.7 d on guinea pigs (n = 432). Daily drop-off of engorged larvae reached peaks on days 7 and 15 from guinea pigs and tortoises, respectively (Fig. 3). The feeding pattern was similar for larvae on both hosts: a phase of slow gain in weight (days 1—4 on guinea pigs, days 1-9 on tortoises) was followed by a phase of rapid engorgement and weight gain before dropping off the host (Fig. 4). The mean drop-off weight of larvae fed on guinea pigs was 2.64 ± 0.22 mg compared with 2.55 ± 0.22 mg for larvae fed on tortoises. The molting period of the engorged larvae to nymphs fed on guinea pigs was 25.9 ± 3.0 d (n = 239) and that of larvae fed on tortoises was 22.1 ± 2.8 d (n = 340).
A 6 8 10 12 2 Days after attachment Fig. 4. Changes in weight (mean ± SD) of larvae of A. marmoreum during their feeding period (based on samples of 10 larvae each). —, tortoise; —, guinea pig.
Feeding Period and Weight of Nymphs. The average feeding period of nymphs was 55.0 ± 15.3 d (n = 76) on tortoises and 7.7 ± 4.3 d (n = 195) on guinea pigs. Peak drop-off from guinea pigs occurred 8 d after attachment, then rapidly declined (Fig. 5). Drop-off from tortoises started 18 d after attachment and continued until day 77 (Fig. 5). No peak drop-off period was evident. The pattern of feeding for nymphs on guinea pigs (Fig. 6) was similar to that of larvae with a period of slight gain in weight (days 1-5), followed by a phase of rapid weight gain which coincided with the start of drop-off of replete nymphs. Feeding on tortoises was also characterized by a period of slow weight gain (Fig. 6). However, the phase of rapid weight gain before repletion could not be detected because engorged nymphs dropped in low numbers (usual8O 7O •
I y/6
8 1O 12 14- 1 6 18 2O 22 24- 26 Days after attachment
Fig. 3. Daily drop-off of A. marmoreum larvae. LTD, tortoise; WM, guinea pig.
6
„n
8 1O 15 25 35 4-5 55 65 75 Days after attachment
Fig. 5. Daily drop-off of A. marmoreum nymphs. LTD, tortoise; • • , guinea pig.
Downloaded from http://jme.oxfordjournals.org/ at University of Nebraska-Lincoln Libraries on June 8, 2016
0
•
.
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