Application of multi-criteria decision analysis to assess sampling strategies in eutrophic urbanized waterbodies.

Different water quality sampling practices such as location selection or frequency can inform future watershed management strategies. The objective of this work was to compare water quality sampling strategies based on different weighted criteria to determine the optimal sampling frequency and sampling location for an urbanized, eutrophic, freshwater system. Weekly water sampling was conducted over a 2-year period at five locations for six water quality parameters. This high frequency (HF) dataset was then deconstructed into a lower frequency (LF) dataset to simulate a monthly sampling strategy. Statistical analyses conducted showed that for all sampling locations the LF datasets were not significantly different from the HF datasets, suggesting monthly sampling is sufficient to capture the overall water quality conditions in this system. A multi-criteria decision analysis was constructed for statistical and operational criteria to determine the optimal sampling locations given different criteria weights. Results showed that the optimal sampling location changed depending on the criteria weighting, suggesting that statistical analyses alone would not be sufficient to determine optimal sampling locations in this system. This analysis was then used if optimal sampling location depended on specific water quality monitoring goals. Results showed that the optimal location depends on the particular water quality monitoring goals and that this effect should also be considered in the design of future sampling programs.

Larval Distribution and Behavior of Chrysomya rufifacies (Macquart) (Diptera: Calliphoridae) Relative to Other Species on Florida Black Bear (Carnivora: Ursidae) Decomposing Carcasses.

Larval interactions of dipteran species, blow flies in particular, were observed and documented daily over time and location on five black bear carcasses in Gainesville, FL, USA, from June 2002 - September 2004. Cochliomyia macellaria (Fabricius) or Chrysomya megacephala (Fabricius) larvae were collected first, after which Chrysomya rufifacies (Macquart) oviposited on the carcasses in multiple locations (i.e., neck, anus, and exposed flesh) not inhabited already by the other blow fly larvae. Within the first week of decomposition, C. rufifacies larvae grew to ≥12 mm, filling the carcasses with thousands of larvae and replacing the other calliphorid larvae either through successful food source competition or by predation. As a result, C. macellaria and C. megacephala were not collected past their third instar feeding stage. The blow fly species, C. megacephala, C. macellaria, Lucilia caeruleiviridis (Macquart), Phormia regina (Meigen), Lucilia sericata (Meigen), and C. rufifacies, completed two developmental cycles in the 88.5-kg carcass. This phenomenon might serve to complicate or prevent the calculation of an accurate postmortem interval.

Hepatozoon species of the timber rattlesnake in northern Florida: description of a new species, evidence of salivary gland oocysts, and a natural cross-familial transmission of an Hepatozoon species.

Two species of Hepatozoon, i.e., H. sauritus and H. horridus n. sp., were present in 1 of 8 timber rattlesnakes, Crotalus horridus. The narrow gamonts of H. sauritus are 15.0-19.0 x 3.5-5.0 microm, with LW 58-86 microm2 and L/W 3.2-4.7, with a narrow, rounded anterior end. The spherical to slightly ovoid oocysts produce ovoid to elongate sporocysts, 21-43 x 12-24 microm, L/W 1.20-2.7, containing on average 22.1 (10-34) sporozoites. This is the first report of a natural cross-familial transfer of a Hepatozoon species. Gamonts of H. horridus n. sp. are 13.0-17.0 x 4.0-6.0 microm, with LW 63-102 microm2 and L/W 2.6-4.0, and have broadly rounded ends. The gamont cytoplasm is vacuolated. The spherical to ovoid oocysts form spherical to elongate sporocysts 14-45 x 11-25 microm, L/W 1.0-2.3, producing an average of 13.0 (8-21) sporozoites. The salivary gland in 1 of 5 mosquitoes dissected contained 1 mature oocyst.

Response of phlebotomine sand flies to light-emitting diode-modified light traps in southern Egypt.

Centers For Disease Control and Prevention (CDC) light traps were modified for use with light-emitting diodes (LED) and compared against a control trap (incandescent light) to determine the effectiveness of blue, green, and red lights against standard incandescent light routinely used for sand fly surveillance. Light traps were baited with dry ice and rotated through a 4 x 4 Latin square design during May, June, and July, 2006. Trapping over 12 trap nights yielded a total of 2,298 sand flies in the village of Bahrif, 6 km north of Aswan on the east bank of the Nile River in southern Egypt. Phlebotomus papatasi comprised 94.4% of trap collections with five other species collected in small numbers. Over half (55.13%) of all sand flies were collected from red light traps and significantly more sand flies (P < 0.05) were collected from red light traps than from blue, green, or incandescent light traps. Red light traps collected more than twice as many sand flies as control (incandescent) traps and > 4 x more than blue and green light traps. Results indicate that LED red light is a more effective substitute for standard incandescent light when surveying in areas where P. papatasi is the predominant sand fly species. Each LED uses approximately 15% of the energy that a standard CDC lamp consumes, extending battery life and effective operating time of traps. Our prototype LED-modified traps performed well in this hot, arid environment with no trap failures.

Two additional Hepatozoon species (Apicomplexa: Hepatozoidae) from the southern black racer, Coluber constrictor priapus (Serpentes: Colubridae), in northern Florida.

Hepatozoon priapus n. sp. from Coluber constrictor priapus has robust gamonts with broadly rounded ends, 18.0 x 4.2 microm (17.0-20.0 x 3.5-6.0), with LW 76.4 microm2 (59-105) and L/W 4.31 (2.9-5.4). The nucleus is always present in second quarter of gamont, seldom extend into first quarter but often into third quarter, 6.0 x 3.0 (5.0-7.0 x 2.5-4.0), with LW 17.9 (13.7-21.0). Erythrocyte cytoplasm is always thin, appearing dehemoglobinized, with infected cells always distorted. Infected erythrocytes are much longer and wider than uninfected cells, with longer nuclei. Oocysts are spherical to ovoid, 92.5 x 86.0 (55-123 x 47-115) and L/W 1.08 (1.0-1.3), contain 14.0 (6-31) sporocysts. Sporocysts, which are also spherical to ovoid, 26.3 x 23.3 (19-50 x 16-38), LW 641.2 (320-1,500) and L/W 1.13 (1.0-2.2), contain 12.6 (5-18) sporozoites. Hepatozoon confusus n. sp., also from C. constrictor priapus, has slender gamonts with rounded ends, 15.6 x 4.1 (14.0-17.0 x 3.5-5.0), with LW 64.3 (52-80) and L/W 3.82 (2.8-4.4). The nucleus is always present in second quarter of gamont, commonly extending into first and third quarters, 5.0 x 2.7 (2.5-4.4 x 4.0-6.0), with LW 13.5 (11.0-16.5). Erythrocyte cytoplasm is sometimes thin, appearing partially dehemoglobinized, with infected cells usually distorted. Infected erythrocytes are longer than uninfected cells but similar in width, with erythrocyte nuclei longer. Oocysts are spherical to ovoid, 115.5 x 108.9 (52-278 x 50-278), with L/W 1.06 (1.0-1.2), and contain 25.0 (7-111) sporocysts. Sporocysts are spherical to ovoid, 27.6 x 25.2 (21-38 x 20-33), LW 701.3 (420-1,125) and L/W 1.09 (1.0-1.4), containing 20.2 (12-32) sporozoites.

Hepatozoon polytopis n. sp. parasitic in two genera and species of colubrid snakes in southern Florida.

Hepatozoon polytopis, described from Coluber constrictor priapus from Palm Beach County, Florida, has short, usually broad gamonts 12.8 x 4.6 microm (10.0-15.0 x 3.5-6.0), with LW 58.5 microm2 (42-84) and L/W 2.84 (1.8-3.7). Nuclei commonly extend into first quarter of gamont (45%), are always present in second quarter, and seldom in third quarter (11%), with dimensions 4.5 x 3.4 (3.0-6.0 x 2.5-4.5) and LW 15.1 (10.0-24.0). Spherical to ovoid oocysts, 122.1 x 104.9 (62-240 x 57-190), with L/W 1.17 (1.0-1.9), contain 31.3 (3-103) sporocysts. Spherical to ovoid sporocysts, 38.0 x 33.9 (28-73 x 25-58), with LW 1,325.1 (756-4,168) and L/W 1.12 (1.0-1.4), contain 42.9 (22-64) sporozoites. Thamnophis sauritus sackenii from Palm Beach County is infected also by H. polytopis, as indicated from similar gamont dimensions and verified by isolation of an identical haplotype of the 18S ribosomal RNA gene from both host species.

Hepatozoon sauritus: a polytopic hemogregarine of three genera and four species of snakes in north Florida, with specific identity verified from genome analysis.

Hemogregarines from Thamnophis s. sirtalis, Coluber constrictor priapus, Elaphe obsoleta quadrivittata, and E. g. guttara in northern Florida appeared to be conspecific on the basis of similar gamonts from all the hosts and sporogonic stages obtained from 3 hosts. The resemblance of gamonts to those of Hepatozoon sauritus, described from T. sauritus sackenii in southern Florida, justified comparison of DNA isolates from the type infection of H. sauritus with samples from each of the northern Florida hosts and with a morphologically distinct species, H. sirtalis, from northern Florida. A nucleotide sequence (530 bp) alignment of the 18S ribosomal RNA gene revealed 2 hemogregarine haplotypes that varied at 15 sites (p distance = 2.8%), which included 10 transitions and 5 transversions. Two well-supported clusters (100% bootstrap support) were revealed by a neighbor-joining tree topology. One cluster included the type infection of H. sauritus and all 4 of the other samples from the northern Florida hosts, with samples of H. sirtalis comprising a second cluster. Hepatozoon sauritus, therefore, is a polytopic species in contrast to the 8 other Hepatozoon species thus far described from snakes in Florida, each of which appears to parasitize a single host species.

Hepatozoon species (Apicomplexa: Hepatozoidae) of the corn snake, Elaphe guttata (Serpentes: Colubridae) and the pigmy rattlesnake, Sistrurus miliarius barbouri (Serpentes: Viperidae) in south Florida.

Hepatozoon guttata n. sp. is described from the corn snake (Elaphe guttata guttata) from south Florida. Gamonts average 14.6 x 4.6 (13-17 x 3.5-6) and are not recurved, with nucleus situated in the second quarter of the gamont. Erythrocyte cytoplasm rarely appears partially dehemoglobinized. The infected cells are usually distorted and are longer than the uninfected erythrocytes but do not differ in width; nuclei are smaller in length and width than those of uninfected cells. Sporogony in Aedes aegypti occurs within the head and the thorax but rarely in the abdomen. Oocysts are spherical to usually ovoid, 87.2 x 75.9 (45-155 x 40-152.5) and contain on average 7.1 (2-32) sporocysts. Sporocysts are spherical to ovoid, 34.8 x 31.0 (20-55 x 17.5-47.5), with 45.7 (14-89) sporozoites contained within. The pigmy rattlesnake (Sistrurus miliarius barbouri) in south Florida is parasitized by H. sistruri n. sp. Gamonts average 14.1 x 5.6 (12.6-15.8 x 4.7-6.3) in breadth and are not recurved, with the nucleus always situated in the second quarter of the gamont commonly at the midbody. Erythrocyte cytoplasm is not dehemoglobinized. The infected cells are always distorted and similar in length to the uninfected cells but with greater width and enlarged nuclei. Sporogony in A. aegypti occurs within head, thorax, and abdomen. Oocysts are spherical to usually ovoid, 163.6 x 154.7 (92-245 x 82-240) and contain 27.4 (12-42) sporocysts. Sporocysts are round to usually ovoid, 39.7 x 33.5 (25-50 x 20-50), with 45.7 (19-70) sporozoites.

The status of Haemogregarina mansoni Sambon and Seligmann from Zamenis flagelliformis Laurenti.

Haemogregarina mansoni Sambon and Seligmann 1907, a parasite of the coachwhip snake (Masticophis flagellum) is redescribed on the basis of sporogonic stages obtained in the experimental vector Aedes aegypti and designated as Hepatozoon mansoni from hosts collected in north Florida. Gamonts average 14.7 x 4.7 (13-16 x 4-6) and are not recurved, with nuclei situated in the second quarter of the gamont. Erythrocyte cytoplasm is often thin, appearing partially dehemoglobinized, or contracted into a central mass with infected erythrocytes always distorted, longer, and more slender than uninfected cells. Sporogony occurred within the head and thorax of A. aegypti. Oocysts were spherical to ovoid, 144.2 x 126.1 (79-198 x 69-178), containing on average 29.2 (7-64) sporocysts. Sporocysts were spherical to ovoid, 33.1 x 29.8 (19-48 x 18-44), with 20.2 (12-32) sporozoites contained within. Experimental infection in Tantilla relicta produced gamonts that did not differ from those in M. flagellum, but dehemoglobinization and cytoplasmic contraction of the host erythrocyte did not occur, and persistent merogonic stages were not present in the tissues.

Haemogregarine specificity in two communities of Florida snakes, with descriptions of six new species of Hepatozoon (Apicomplexa: Hepatozoidae) and a possible species of Haemogregarina (Apicomplexa: Haemogregarinidae).

Five species of snakes in Florida, from Palm Beach County in the south and Alachua County 450 km to the north, occur in similar habitat but have distinctive Hepatozoon species characteristic of each host species. In Palm Beach County, Diadophis punctatus is host to Hepatozoon punctatus n. sp., Thamnophis sauritus sackenii to Hepatozoon sauritus n. sp., and Nerodia fasciata pictiventris to Hepatozoon pictiventris n. sp. In Alachua County, N. fasciata pictiventris is parasitized by Hepatozoon fasciatae n. sp., Seminatrix p. pygaea by Hepatozoon seminatrici n. sp., and Thamnophis s. sirtalis by Hepatozoon sirtalis n. sp. Each Hepatozoon sp. has distinctive gamonts and sporogonic characters and, in the 4 species where known, meronts. Nerodia floridana is host to Haemogregarina floridana n. sp. in both localities, with generic identification tentative, based upon presence of erythrocytic meronts. The presence of sporocysts in the proboscis of 31% of Aedes aegypti infected by H. pictiventris is the first report of infective stages of a reptilian Hepatozoon species within the mouthparts of a dipteran vector. This study suggests that in Florida, at least, the diversity of the Hepatozoon community not only equals but probably exceeds the diversity of the snake communities present, and that host specificity in nature may be much greater than that postulated from previous studies.

Effects of temperature on Sarcophaga haemorrhoidalis (Diptera: Sarcophagidae) development.

Growth curves were studied for the larva and pupa of Sarcophaga haemorrhoidalis (Fallen) under cyclic temperatures with means of 15.6, 21.1, 26.7, and 35 degrees C, and a constant temperature of 25 degrees C. Development from 1st instar to adult under all regimes ranged from 252 to 802 h. A constant temperature of 25 degrees C produced a range of pupation times from 93 to 153 h, with adult emergence ranging from 252 to 502 h. A maximal preferential temperature of 30 degrees C was determined for maggots by using a gradient system. This study shows that S. haemorrhoidalis can be used as an indicator species when establishing a postmortem interval as long as the investigator accounts for the developmental variation prevalent within this species.

Field evaluation of colored light-emitting diodes as attractants for woodland mosquitoes and other diptera in north central Florida.

The attraction of mosquitoes to transmitted light from colored super-bright light-emitting diodes (LEDs) (100-nm bandwidth) was evaluated by comparison of capture numbers with and without carbon dioxide-baited (200 ml/min) Centers for Disease Control (CDC) traps. Traps with either colored LEDs or control lights were arranged in Latin square designs at 2 north central Florida woodland locations and checked daily during July and August 1996. When data were analyzed by species, a significant difference in attractivity of lights was found in some species. Aedes atlanticus, Aedes dupreei, Aedes infirmatus, Anopheles crucians s.l., Culiseta melanura, Culex nigripalpus, Psorophora columbiae, and Uranotaenia sapphirina showed significant color preferences. These results will have potential for use by ecologists, epidemiologists, and mosquito control personnel for improving collection efficiency of certain species of mosquitoes.

Granulomatous hepatitis associated with Hepatozoon sp. meronts in a southern water snake (Nerodia fasciata pictiventris).

A wild-caught adult female southern water snake (Nerodia fasciata pictiventris) did poorly in captivity. A peripheral blood-film examination demonstrated numerous hemogregarines characterized as fusiform nondividing intraerythrocytic gametocytes. Xenodiagnostic typing in laboratory-reared mosquitoes demonstrated the parasite to be of the genus Hepatozoon. Gross and histopathologic examination of the liver demonstrated numerous granulomas centered on groups of one to six Hepatozoon sp. meronts, an unusual finding in naturally infected wild-caught snakes.

Artificial feeding of Amblyomma cajennense (Fabricius, 1787) (Acari:Ixodidae) through silicone membrane.

An artificial feeding system was used where citrated bovine blood was offered to male and female Amblyomma cajennense. Vestiges of blood, sweat, hair and exfoliated skin were used as phago-stimulants placed on the surface of the silicone membrane. The ticks were collected, as engorged nymphs, from naturally infested equines, with the ecdysis occurring in the laboratory. Four hundred ticks were used, 50% being female, at three to four weeks post-ecdysis. Vestiges of blood on the silicone membrane were the most efficient phago-stimulant and the association of vestiges of blood and sweat residue smears yielded better results compared to the other phago-stimulants used.

Effects of temperature on Chrysomya rufifacies (Diptera:Calliphoridae) development.

Growth curves were studied for the egg, larva, and pupa of Chrysomya rufifacies (Macquart) under mean cyclic temperatures of 15.6, 21.1, 26.7, and 35.0 degrees C and a constant temperature of 25.0 degrees C. Development from egg to adult under all regimes ranged from 190 to 598 h. A constant temperature of 25 degrees C produced a range of pupation times from 134 to 162 h, with adult emergence ranging from 237 to 289 h. The maximal preferential temperature of 35.1 degrees C was determined for maggots using a gradient system. Highly predictable developmental time, minimal larval length variation, and low cohort variation emphasize the utility of this species in entomological-based postmortem interval determinations. Therefore, C. rufifacies should be of primary forensic importance when recovered alone or in conjunction with other species of Calliphoridae.

Effects of temperature on Cochliomyia macellaria (Diptera:Calliphoridae) development.

Growth curves were studied for the egg, larva, and pupa of the secondary screwworm, Cochliomyia macellaria (F), under mean cyclic temperatures of 15.6, 21.1, 26.7, 32.2, and a constant 25.0 degrees C. Development from egg to adult under all regimes ranged from 170 to 588 h. A constant temperature regime of 25 degrees C produced a range of pupation times from 92 to 142 h, with adult emergence ranging from 196 to 261 h. The maximal preferential temperature of 39 degrees C was determined for maggots by using a gradient system. The minimal variation in the range of larval lengths and development time makes this species suitable for establishing relatively narrow confidence intervals when constructing a postmortem interval estimate for use in legal investigations.

Immune recognition of Ornithodoros tick (Acari: Argasidae) salivary antigens by anti-Psoroptes cuniculi antibodies.

Rabbits infested with either Ornithodoros sp. ticks or Psoroptes cuniculi mites were assayed for anti-tick antibody by enzyme-linked immunosorbent assay. Titration of rabbit serum against Ornithodoros sp. salivary gland extract (SGE) demonstrated both mite- and tick-infested animals to have elevated anti-tick antibody titers. Western blot analysis demonstrated the anti-mite and anti-tick antisera to contain antibodies with affinities for both common and unique subsets of Ornithodoros SGE proteins.

Isolation and characterization of mouse monoclonal antibodies reactive with soft tick (Acari: Argasidae) salivary antigens of high molecular weight.

The purpose of this study was to immunologically characterize soft tick salivary antigens. BALB/c mice hyperimmunized with salivary gland extract prepared from Ornithodoros talaje (Guérin-Méneville) were observed to develop high titers of antitick salivary antigen antibodies. Subsequent fusion of splenic lymphocytes from the hyperimmunized mice with SP-2/0-AG14 myeloma cells resulted in the production of 10 antitick IgM-producing hybridoma clones. Partial characterization of the respective tick antigens by gel filtration and SDS-PAGE demonstrated all 10 monoclonal antitick antibodies to be reactive with a salivary gland extract fraction containing proteins 50-110 kDa in molecular weight. Cross-reactivity assays and electrophoretic comparison of salivary gland extract specimens demonstrated similar proteins in several ixodid tick genera and species.

Detection and quantification of Ornithodoros-specific anti-tick antibody by competitive inhibition ELISA.

The objective of this study was to develop a highly specific enzyme-linked immunosorbent assay (ELISA) for the serological detection of anti-Ornithodoros tick antibodies in animals. Affinity-purified rabbit anti-Ornithodoros IgG antibodies were employed in indirect competitive inhibition ELISA assays designed to measure the anti-Ornithodoros antibody titers in other animal species using the domestic goat (Capra hircus) as a large animal model. Repeated infestation of goats with Ornithodoros coriaceus was found to elicit the formation of antibodies capable of inhibiting the binding of the Ornithodoros-specific rabbit IgG. Western blot analysis of goat and rabbit anti-tick antisera demonstrated both animal species to respond immunologically to a set of 9 major protein bands in O. coriaceus salivary gland extracts. The results of these experiments demonstrate that a history of animal exposure to O. coriaceus may be detected serologically by competitive inhibition ELISA.