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In a two-lactation study, 352 Holstein cows (124 primiparous) from six commercial dairy herds were assigned to daily injections of bST. Dosages were 0 (control), 5.15, 10.3, or 16.5 mg/d of bST; injections began 28 to 35 d postpartum. During yr 1, FCM production increased linearly as bST dose increased for primiparous and multiparous cows. However, FCM production for primiparous cows that were retained for yr 2 was not increased by bST, and the increase in FCM for multiparous cows was only 67% of that observed during yr 1 for doses of 10.3 and 16.5 mg/d. Milk SCC were not increased with the use of bST during either lactation. Days to conception of multiparous cows during yr 1 tended to increase linearly with increasing dose. Multiparous cows administered bST during yr 2 tended to have a lower conception rate than control cows (71 vs. 87%, respectively). Body condition was decreased linearly by bST dose at the end of yr 1 and 2. Prior to receiving bST during yr 2, bST-treated cows regained less body condition than control cows. Number of health disorders at parturition and assisted births of cows that received bST during yr 1 were unaffected by previous treatment. General health, ratio of single to multiple births, and BW gain of calves was similar among treatments.

Leptospira interrogans serovar grippotyphosa was isolated from the skin of a 14-year-old male dog with deteriorating health. Necropsy revealed numerous lesions characteristic of aged dogs, but no evidence of acute hepatitis or nephritis, which are common features of pathogenic Leptospira infections. Antibody to Leptospira was not detected in the dog's serum by microagglutination. Leptospires grew slowly in Barbour-Stoenner-Kelly medium, a medium commonly used to isolate Borrelia, but then grew abundantly in Tween 80-bovine albumin leptospire medium. The isolate was pathogenic to a hamster and was identified by microagglutination and restriction endonuclease analysis.

The genomes of 62 North American and European Borrelia burgdorferi isolates were examined by restriction endonuclease analysis (REA), gene probe restriction fragment length polymorphism, and pulsed-field gel electrophoresis (PFGE). Hybridization of restriction fragments with the immunologically relevant 83-kDa antigen gene revealed polymorphisms and divided the isolates into three major groups. Group I included all but two of the American isolates and some of the European isolates. One of two Californian isolates (DN 127) and one Ixodes dammini isolate from New York (strain 25015), previously described as atypical, were distinct from the isolates in the three groups. Plasmid profile analysis and REA, the method with the highest level of discrimination, revealed extensive heterogeneity among isolates of the same major group. Our study demonstrates the usefulness of the 83-kDa antigen gene probe for dividing the isolates into major genogroups, whereas REA and plasmid profile analysis allow for a distinction of individual strains within these groups.

In a retrospective study, indirect fluorescent-antibody staining methods were used to detect immunoglobulins to Ehrlichia canis and Ehrlichia risticii in canine and equine sera that had originally been analyzed for antibodies to Borrelia burgdorferi. Analyses of 60 dog serum specimens collected in Connecticut and New York State during 1986 revealed antibodies to E. canis in 7 (11.7%) specimens; titration endpoints ranged from 1:40 to 1:320. Three of these dogs had anemia. Of the 187 equine serum specimens obtained in Connecticut during 1985 and analyzed by indirect fluorescent-antibody staining methods, 17 (9.1%) contained antibodies to E. risticii. Maximal antibody titers of 1:1,280 were recorded for serum specimens collected from three equids during May and July. We conclude that canine and equine ehrlichiosis coexist with Lyme borreliosis in Connecticut and the lower Hudson River Valley of New York State.

Enzyme-linked immunosorbent assays (ELISA) and immunoblots using either whole-cell lysates of Borrelia burgdorferi or an antigenic region of flagellin (41-G) as the antigen were performed, and the abilities of the two assays to detect antibodies to this spirochete in dog, cottontail rabbit, and horse sera were compared. Assays using whole-cell B. burgdorferi lysates as the antigen were more sensitive for detecting antibodies. ELISA with 41-G as the antigen were specific for Borrelia antibodies but were not as sensitive as the assays with whole-cell lysates coated to the solid phase. Use of recombinant full-length flagellin, rather than 41-G, as the antigen in immunoblots increased the sensitivity of each assay. However, antibodies to other bacterial antigens cross-react with whole flagellin and may account for false-positive results. Antibodies to B. burgdorferi outer surface protein A or B were usually undetected when the sera were tested by immunoblotting methods. Borrelia lysates or the 41-G antigen may be used in ELISA or immunoblots to document host exposure to this spirochete. The use of 41-G as the antigen may increase the specificity of an assay or help confirm the serologic diagnosis of Lyme borreliosis in dogs, horses, and cottontail rabbits.

The effect of photoperiod on longevity of nondiapausing members of a species in the mosquito species complex Anopheles quadrimaculatus was tested in laboratory experiments. First-generation adults reared from 2 field collections showed the same trends: those reared under short photoperiod (8 h light: 16 h dark) lived longer than did those reared under long photoperiod (16 h light: 8 h dark). Adult metabolic rates were not consistently related to longevity. In adults reared from the first collection, metabolic rates were higher in long-photoperiod individuals but in adults reared from the second collection, metabolic rates were higher in short-photoperiod individuals. Longevity appears to be another of many insect features that are affected similarly by temperature and photoperiod.

The outer surface proteins OspA and OspB of Borrelia burgdorferi have recently been demonstrated to be major target proteins for protective antibodies in mice against infection with the homologous spirochaetal strain. However, it has become clear from a variety of studies that B. burgdorferi isolates of different geographical origin and/or sources are heterogeneous and that they can be divided into at least six subgroups according to their distinct OspA/OspB genotypes. In order to analyse cross-protection between these subgroups we have now generated immune sera to various isolates of B. burgdorferi with different OspA/OspB genotypes. We show that passive immunization with antisera specific for whole spirochaetes or recombinant OspA of one spirochaetal isolate protects severe combined immunodeficiency mice against infection with strains of the corresponding OspA/OspB genotype but only partially or not at all against infection with isolates expressing distinct OspA/OspB genotypes. The incomplete protection mediated by individual antisera against independent isolates of B. burgdorferi suggests that an effective subunit vaccine against Lyme disease should consist of a mixture of OspA structures covering the heterogeneity of this protein within the species B. burgdorferi.

Ticks and blood specimens were collected from white-tailed deer (Odocoileus virginianus) in Connecticut and analyzed to identify foci for Lyme borreliosis. Males and females of Ixodes scapularis, the chief vector of Borrelia burgdorferi, were collected from deer in five of eight counties during 1989-1991. Analysis by indirect fluorescent antibody (IFA) staining of midgut tissues showed that prevalence of infection was highest (9.5% of 367 ticks) in south central and southeastern Connecticut. Infected I. scapularis also were collected from southwestern regions of the state (12.1% of 99 ticks), but prevalence of infection in northern counties was considerably lower (0.8% of 124 ticks). Deer sera, obtained in 1980 and 1989-1991, were analyzed by an enzyme-linked immunosorbent assay or by IFA staining methods. Antibodies to B. burgdorferi were detected in sera collected from all eight counties in Connecticut. Deer had been infected by this spirochete in at least 50 towns, 17 (34%) of which are in south central and southeastern parts of the state. Borrelia burgdorferi is widely distributed in I. scapularis populations in Connecticut.

A recombinant protein (p41-G) of an antigenic region of flagellin was used in a standard and amplified enzyme-linked immunosorbent assay (ELISA) to detect antibodies to Borrelia burgdorferi, the causative agent of Lyme borreliosis. Comparable sensitivities (88 to 94%) were noted when sera from 17 persons who had erythema migrans and antibodies to whole-cell B. burgdorferi were tested against the p41-G antigen. In tests of a second study group of 36 persons who had erythema migrans but no detectable antibodies to whole-cell B. burgdorferi, 3 (8%) were positive when the p41-G antigen was used. Assay specificity likewise increased when the p41-G fragment was included in an ELISA with human sera containing treponemal antibodies. Recombinant flagellar proteins of B. burgdorferi, such as the p41-G antigen, can be used in an ELISA and may help confirm Lyme borreliosis during early stages of infection and improve specificity.

Lyme disease is prevalent over vast areas of land in the northern hemisphere. The etiologic agent is a spirochete, Borrelia burgdorferi, that is transmitted by Ixodes ticks. At least five species of anthropophilic ticks are competent vectors, and the causative agent or closely related species have been detected in 20 wild or domestic mammals and eight birds. The ability to isolate and culture B. burgdorferi from humans, ticks, and wildlife has significantly contributed to our understanding of the epizootiology of Lyme disease. Specific procedures for isolating borreliae from ticks and rodents in Barbour-Stoenner-Kelly medium are described. The authors have preserved in liquid nitrogen more than 500 isolates from ticks, wildlife, and a human.

Serologic studies were conducted to determine whether white-footed mice (Peromyscus leucopus) and cotton mice (Peromyscus gossypinus) contained serum antibodies to Borrelia burgdorferi, the causative agent of Lyme borreliosis. Enzyme-linked immunosorbent assays detected antibodies to this spirochete in 35.7 and 27.3% of 56 P. leucopus and 535 P. gossypinus serum samples, respectively, collected in Connecticut, North Carolina, South Carolina, Georgia, Florida, Alabama, and Mississippi. Antibody titers ranged from 1:160 to greater than or equal to 1:40,960. On the basis of adsorption tests, the antibodies detected appeared to be specific to Borrelia spirochetes. Seropositive rodents in the eastern and southern United States, areas where human cases of Lyme borreliosis have been reported, indicate a widespread geographic distribution of B. burgdorferi or a closely related spirochete.

The objective of this study was to determine whether different isolates of Babesia microti could be distinguished from morphologically similar isolates of B. gibsoni by using a ribosomal DNA (rDNA) probe. A Babesia-specific rDNA probe was obtained by polymerase chain reaction amplification of sequences from B. microti DNA using universal primers directed against highly conserved portions of the eukaryotic 16S-like rRNA gene. The chemiluminescent rDNA probe hybridized to Southern blots of restriction endonuclease-digested DNA preparations of different isolates of B. gibsoni from infected dogs and B. microti from infected humans and white-footed mice. Restriction fragment length polymorphisms served to differentiate these species. Although the hybridization patterns seen with DNAs from six B. microti isolates did not vary, those of the five B. gibsoni isolates did indicate genotypic variation. We concluded that isolates of B. microti and B. gibsoni can be differentiated on the basis of restriction fragment length polymorphism detected with a chemiluminescent rDNA probe.

Babesia microti was isolated from a white-footed mouse (Peromyscus leucopus) that was captured in southeastern Connecticut in 1988, when the first human case of babesiosis acquired in Connecticut was recognized. To date, 13 cases of babesiosis have been reported in Connecticut, the largest number of human cases reported on the mainland United States. Two of nine patients quiried remembered a prior tick bite. Since Babesia parasites are known to be vectored only by ticks, we surmise that 12 of these infections were acquired via tick bites; 1 was obtained by blood transfusion (the patient was 46 years of age) from an endemically infected donor. The ages of the patients with tick-acquired babesiosis ranged from 61 to 95 years. Two patients died with active infections, and one patient died from chronic obstructive pulmonary disease soon after treatment with clindamycin and quinine. Indirect fluorescent-antibody titers of blood samples drawn at the time of hospitalization for 11 patients and at the time of active infection for 1 asymptomatic person ranged from 1:1,024 to 1:4,096. Five of eight patients with babesiosis also had significant immunoglobulin G or immunoglobulin M titers (1:640 to 1:5,120) to Borrelia burgdorferi. B. microti was isolated in Syrian hamsters inoculated with blood from 7 of 12 patients tested and was also isolated from mice captured in six towns. The peridomestic nature of the disease was demonstrated by isolating the parasite from white-footed mice captured in or near the yards of eight different patients. Of 59 mice tested, 27 were positive and 25 were coinfected with B. burgdorferi. The isolation of B. microti from a white-footed mouse captured in north-central Connecticut (West Hartford), away from the focus of human infections in southeastern Connecticut, suggests that this pathogen may spread into other areas where Ixodes dammini, the tick vector, becomes established.

An enzyme-linked immunosorbent assay (ELISA) was developed to detect serum antibodies to Borrelia burgdorferi, the causative agent of Lyme borreliosis, in deer (Odocoileus virginianus) and raccoons (Procyon lotor). Blood samples were collected from these mammals in Connecticut, Maryland, North Carolina, Georgia and Florida. Seropositivity for deer was highest in Connecticut (56% of 353 sera) and Maryland (51% of 35 sera). Raccoons in Connecticut, Maryland, North Carolina, and Florida also had antibodies to B. burgdorferi, but prevalence of positive sera was highest in Maryland (79% of 14 samples). Based on adsorption tests, the immunoglobulins detected in these mammals were probably specific to B. burgdorferi. The ELISA was more sensitive than an indirect fluorescent antibody staining method and was more suitable for analyzing large numbers of serum samples.

Serum samples from persons with Lyme borreliosis, periodontitis, or acute necrotizing ulcerative gingivitis were analyzed by an enzyme-linked immunosorbent assay (ELISA) with and without adsorption and amplification procedures. When biotin and streptavidin reagents were used as an amplification procedure in ELISA without the use of commercially prepared sorbent (Treponema phagedenis biotype Reiter), sensitivity increased. Of the 85 serum samples collected from persons with erythema migrans but no detectable antibodies to Borrelia burgdorferi by standard ELISA, 17 (20%) were reactive after amplification. Adsorption of serum samples with a 1:10 dilution of T. phagedenis biotype Reiter sorbent used in conjunction with amplified ELISA also improved the sensitivity of this method. However, cross-reactivity could not be completely eliminated. An adsorbed-amplified ELISA may be helpful in the diagnosis of Lyme borreliosis in the laboratory, particularly during early weeks of infection, when antibodies to B. burgdorferi can be present at a low concentration.

Ixodes dammini ticks from two northwestern Illinois sites were found to be infected with Borrelia burgdorferi at rates of 19 and 32%. B. burgdorferi isolates, one from each site, had protein and antigenic patterns similar to those of the B-31 strain. An indirect immunofluorescence method proved to be more sensitive than dark-field microscopy in detection of these spirochetes. A modified BSK medium containing rifampin was found to be more efficient for spirochete isolation than unsupplemented BSK medium.

An increasing number of phenotypically heterogeneous borrelia isolates have been obtained by culture. Genetic analysis of some of them suggest that a reorganisation in delineation of the different species within the genus Borrelia may be necessary. Specifically increasing numbers of variants phenotypically different from the American type strain of B. burgdorferi have been described. In this workshop experts have discussed phenotypic and genotypic characters for (a) differentiation of B. burgdorferi strains from other borrelia and from treponemes, (b) characterization of B. burgdorferi isolates at the subspecies level. Ultrastructure was discussed by K. Hovind-Hougen, antigen structure by B. Wilske and genotypic characters by G. Baranton, A. G. Barbour, R. C. Johnson and V. Preac-Mursic. J. F. Anderson discussed differences between strains isolated from a wide range of vectors and hosts in North America. The outcome of the workshop was that B. burgdorferi may comprise different genomic species which however share common epitopes recognized by certain monoclonal antibodies. At the subspecies level a broad heterogeneity was demonstrated using methods as restriction endonuclease analysis, hybridization with whole B. burgdorferi-DNA or specific probes as well as plasmid analysis. A serotyping system based on monoclonal antibody reactivity against the outer surface protein OspA was proposed. A comparison of the different typing methods is impaired by use of different strains for analysis. In the future a broad variety of phenotypically different and defined strains need to be analysed for genotypic clusters and later on reexamined for phenotypic characters. To accomplish this close cooperation of different research groups is necessary.

1. Descendents of a Florida, U.S.A. population of Drosophila melanogaster were reared under short (8 hr light: 16 hr dark) and long (16 hr light: 8 hr dark) photoperiods. 2. Flies reared under short photoperiods had higher rates of metabolism at 21 degrees C. 3. Because genetic background (iso-female line) affects metabolic rates, statistical control through analysis of covariance was necessary to isolate the effects of photoperiod on metabolic rates. 4. These results on a subtropical population of D. melanogaster are similar to those found on a temperate population of the same species.

At the IV International Conference on Lyme Borreliosis, a workshop was held to identify the unique development of the Lyme disease spirochete, Borrelia burgdorferi, in its established and suspected arthropod vectors. The following is a summary of the panel's discussions of research aspects concerning relationship(s) of this borrelia to its vectors, and the mode(s) of its transmission to animal hosts.

Borrelia burgdorferi is transmitted from animals to humans by ticks belonging primarily to the Ixodes ricinus complex. Transmission occurs during feeding, either by salivation, regurgitation, or by both processes. Two Eurasian (Ixodes ricinus and I. persulcatus) and three North American (I. dammini, I. scapularis and I. pacificus) species vector B. burgdorferi to humans. All ticks feed on three different host animals during their lives, and each species parasitizes a relatively large number of different host animals. For example, Ixodes ricinus is known to feed on more than 300 different kinds of mammals, birds, and reptiles. Borrelia burgdorferi has been isolated or detected in 24 different species of mammals or birds. Rodents belonging to the genera Peromyscus and Apodemus are important reservoirs in North America and Europe, respectively. Birds are a natural means of distributing infected ticks long distances during their migratory flights. Reptiles may not support the growth of borreliae. The B31 strain of B. burgdorferi is predominant in north-eastern United States. It is the only strain known to cause disease in humans, and the only one thus far isolated from rodents. In Europe, variants with distinctly different proteins from those found in the B31 strain also are infectious to humans, through they have not been identified with any specific clinical syndrome of Lyme borreliosis. Variants of the B31 strain have been isolated from lagomorphs in the United States.

We document for the first time an infectious but nonarthritogenic variant of Borrelia burgdorferi. Strain 25015, previously isolated from an Ixodes dammini larva collected in upstate New York, was infectious but failed to produce arthritis or carditis in laboratory rats and mice. By contrast, pathogenic strain N40 invariably caused arthritis. This nonarthritogenic variant, with proteins with molecular weights different from those of the standard B31 strain, was frequently isolated from normal joint tissues of experimentally infected rats. Outer surface proteins A and B of strain 25015 have molecular weights of about 32,500 and 35,500, respectively, in contrast to molecular weights of approximately 31,000 and 34,000, respectively, for outer surface proteins A and B of strains B31 and N40. A prominent low-molecular-weight protein of about 23,500 also characterizes strain 25015. Test animals infected for 30 to 60 days had relatively high antibody titers (greater than or equal to 1:1,280). The nonarthritogenic variant will be useful, along with pathogenic strains, in providing comparative insight into the pathogenesis of Lyme borreliosis. Homologous immunoblotting of sera from rats and mice inoculated with both the arthritogenic and nonarthritogenic strains revealed antibody reactivities to proteins of B. burgdorferi different from those revealed in the heterologous tests.

Ticks were removed from naturally infested cats, and serum samples from these cats were tested for antibodies to Borrelia burgdorferi. Twenty-two of 93 cats (23.7%) had one or more motile stages of Ixodes dammini attached. Of 2 larvae and 20 nymphs removed from cats, 1 larva and 2 nymphs were infected with B burgdorferi. Spirochetes were not found in tissues of 13 female and 4 male ticks. Ten of 71 serum samples analyzed by indirect fluorescent antibody staining or ELISA contained antibodies to this spirochete. Maximal antibody titers were 1:256 and 1:2,560, respectively. At titers greater than or equal to 1:160 in ELISA, seropositivity ranged from 8.8% (n = 34 sera tested from 34 cats) in May through July to 33.3% (n = 12 cats tested) during February through April. In clinical studies of 30 cats, there were nearly equal percentages of seropositive cats with limb or joint disorders not accompanied by fever, anorexia, or fatigue (5 of 21 cats) and cats with these signs of illness but lacking lameness (2 of 9 cats.)

Serum samples obtained from 59 persons who had acute necrotizing ulcerative gingivitis, periodontitis, syphilis, or Lyme disease were tested against Treponema phagedenis biotype Reiter, Treponema denticola, Treponema vincentii, and Treponema scoliodontum by indirect fluorescent-antibody staining methods. Although there were positive reactions for sera representing each of these study groups and for 20 (13%) of 156 samples collected from the general population (premarital screening for syphilis), titration endpoints were relatively low (less than or equal to 1:256). Serum samples from 18 persons who had gingivitis or periodontitis but no history of Lyme borreliosis were tested by enzyme-linked immunosorbent assay for antibodies to Borrelia burgdorferi. Of these, five (28%) had immunoglobulin M antibody and four (22%) contained immunoglobulin G antibodies to this spirochete. Adsorption with either sorbent commercially prepared from T. phagedenis biotype Reiter or with washed, whole cells of T. phagedenis biotype Reiter reduced cross-reactivity in the enzyme-linked immunosorbent assay for Lyme borreliosis.

An enzyme-linked immunosorbent assay was developed to detect antibodies to Borrelia burgdorferi in cottontail rabbits captured in Millbrook, N.Y., and New York, N.Y. Five antigenically variable strains of B. burgdorferi were analyzed to determine the variability of serologic test results. In analyses of 79 serum samples, seropositivity ranged from 56% for a strain cultured from kidney tissues of a cottontail rabbit to 68% for a strain isolated from a larva of Ixodes dentatus, a tick that parasitized a cottontail rabbit. There were false-positive results when reference rabbit antisera to B. hermsii and Treponema pallidum were screened against B. burgdorferi. Cross-reactivity with antisera to Leptospira interrogans serovars was less pronounced. Western blot (immunoblot) analyses revealed reactivities of test sera to two or more surface or subsurface proteins of B. burgdorferi with approximate molecular masses of 18, 25 to 27, 34, 36, 41, and 59 kilodaltons. Cottontail rabbits respond immunologically to B. burgdorferi, but the observed variations in serologic test results should not be a limitation in field and laboratory investigations of Lyme borreliosis.

Multiple blood samples were obtained from privately owned dogs living in tick-infested areas of New York (Westchester County) and Connecticut, where Lyme disease in human beings has been reported. Of the 175 dogs examined, 127 (72.6%) had limb/joint disorder, whereas the remaining 48 dogs were considered healthy. Results of analysis of 419 serum samples revealed IgM antibody to Borrelia burgdorferi in healthy and lame dogs during all seasons. Prevalence of seropositivity was significantly (P less than 0.01) greater, using a polyvalent ELISA (89.5%) than using a class-specific ELISA for IGM antibody (57.8%). Mean antibody titers obtained by use of polyvalent ELISA were likewise higher than IgM titers. Analysis of paired serum samples from dogs with limb/joint disorder indicated that 118 (92.9%) remained positive for IgM or IgG antibodies when retested weeks or months after initial testing. In 48 dogs without history of joint involvement or other signs of disease, 43 (89.6%) had antibody to B burgdorferi 2 or more times. Serotest results also revealed little or no change in antibody titer for lame dogs given antibiotics or for healthy dogs 2 or more months after initial sample collection.

The first case of canine ehrlichiosis in Connecticut is reported. A female Brittany spaniel from Milford presented with lethargy, anorexia, fever, petechiae, splenomegaly, thrombocytopenia, anemia, elevated serum alkaline phosphatase, lymphopenia, and hypoalbuminemia. Serologic analysis revealed antibodies to Ehrlichia canis (titer, 1:2,560). This documents a more northern geographic distribution in the United States for this infectious agent than had previously been suspected.

Bird-feeding Ixodes dammini ticks were documented for the first time to successfully molt and transstadially pass Borrelia burgdorferi spirochetes that were indistinguishable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis from the type B31 strain. Forty-six of 73 blood-engorged larvae and 50 of 66 fully-fed nymphs, removed from wild-caught birds, successfully molted. Borreliae were isolated from 21 of 78 partially- and fully-fed larvae off birds, including six specimens that molted. Spirochete-positive cultures also were obtained from 35 of 60 partially- and fully-fed nymphs that had fed from birds, including 20 nymphs that molted into adult ticks. Transstadially passed borreliae by bird-feeding larval and nymphal I. dammini were infectious to hamsters, leading us to suggest that these ticks are capable of subsequently transmitting infectious spirochetes to mammals, including humans. An isolated of B. burgdorferi, recovered from a bird-feeding larval Ixodes dentatus, was indistinguishable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis from the B31 strain. This isolate, unlike another from I. dentatus off a cottontail rabbit (Sylvilagus floridanus), had a protein band with a molecular weight of approximately 31,000 that reacted with murine monoclonal antibodies H3TS and H5332 in western blot analysis. Thus, closely related borreliae are present in both I. dentatus and I. dammini.

1. Second-generation descendents of wild-caught female Drosophila simulans Sturtevant collected at different seasons from Gainesville, Florida, USA, were reared under a constant temperature and food level but with seasonally variable photoperiods. 2. Body weight and metabolic rate, after adjustment to a common body weight, were higher in descendents of flies collected in fall and winter. 3. Temperature sensitivity (Q10) of metabolic rate was also related to season: higher Q10 values were associated with seasons of increasing temperatures and lower Q10 values were associated with seasons of decreasing temperatures. 4. These metabolic characteristics may be adaptive in enhancing activity at lower temperatures and conserving energy at higher temperatures.

Spirochetes and their vectors and reservoirs were studied in a Lyme disease focus in East Haddam, Connecticut, from mid-May through September 1983. Ixodes dammini subadults were comparable in number on white-footed mice (Peromyscus leucopus) (means = 2.9 +/- 3.6 SD) to those on 27 different species of birds (means = 2.3 +/- 4.2 SD) representing 11 families within the order Passeriformes. Less commonly found ticks on birds (means less than or equal to 0.1) were immature Ixodes dentatus and Haemaphysalis leporispalustris. Although spirochete-infected I. dammini larvae and nymphs were taken off eight and nine different species of birds, respectively, significantly fewer positive larvae were removed from birds than from white-footed mice. Spirochetes were detected in the midguts of I. dammini, Dermacentor variabilis, and H. leporispalustris and two species of insects (Cuterebra fontinella and Orchopeas leucopus). Possibly, arthropods other than I. dammini vector these spirochetes in northeastern United States. Spirochetes grew in a cell-free medium inoculated with bloods from four white-footed mice, one woodland jumping mouse (Napaeozapus insignis), one northern mockingbird (Mimus polyglottos), one gray catbird (Dumetella carolinensis), two prairie warblers (Dendroica discolor), one orchard oriole (Icterus spurius), one common yellowthroat (Geothlypis trichas), and one American robin (Turdus migratorius). We suggest that avian hosts, like mammals, develop spirochetemias of the causative agent of Lyme disease. Erythematous tissues from a white-footed mouse were infected with spirochetes.

We prepared fractions of Borrelia burgdorferi, the etiologic agent of Lyme disease, from cultured spirochetes and used them as antigen in an enzyme-linked immunosorbent assay (ELISA) for IgG antibody. Polystyrene plates coated with an extract containing major proteins with apparent molecular masses of 34, 39, 59, and 68 kilodaltons had comparable sensitivity but greater specificity than plates coated with whole cells. Of the 33 serum specimens from individuals with Lyme disease that reacted with whole cells of B. burgdorferi in the class-specific ELISA, 30 (91%) remained positive when this extract was used. Cross-reactivity was minimal with antibody to treponemes. Use of subunit antigens may improve serological diagnosis of Lyme disease.

A modified membrana nictitans flap technique was used successfully in 1,768 of 1,845 (95.8%) cattle with several types of ulcerative keratitis. The flap was easily applied and lasted 7 to 10 days.

An enzyme-linked immunosorbent assay was compared with an indirect fluorescent antibody test for its ability to detect antibodies to the Lyme disease spirochete in sera of naturally infected humans, dogs, and white-footed mice and experimentally infected Swiss mice. Ninety-five percent of the total 123 sera analyzed reacted similarly in both tests. For 36 human sera, the correlation coefficient (r = 0.47) for logarithmic transformations of indirect fluorescent antibody and enzyme-linked immunosorbent assay titers was significant at P less than 0.01. Within each mammalian species, mean titers for indirect fluorescent antibody and enzyme-linked immunosorbent assay antibodies were within three-fold. Comparisons of different naturally infected mammals revealed relatively higher average titration endpoints in both tests for patients with Lyme disease. Human sera also had the widest range of titers. Both methods proved satisfactory for serological confirmation of prior spirochetal infections.

A species of Babesia which infects raccoons, Procyon lotor, is described and named Babesia lotori. Twelve of 14 raccoons captured from five counties in Connecticut developed parasitemia after splenectomy. Preoperative levels were subpatent or less than 1%. Parasitemia peaked anywhere from less than 5% to 36.6% in splenectomized raccoons, and all but one survived and possessed low infection rates (less than or equal to 3%) at the end of the experiment. An indirect fluorescent antibody test is described and shown to be useful in detecting antibodies to B. lotori. Seropositive raccoons were obtained from six counties. Two, Babesia-free, splenectomized raccoons developed parasitemia with corresponding increases in antibody titers after they were inoculated with infected blood. Antibody titers of naturally infected raccoons following splenectomy either remained stable or increased fourfold. Babesia was observed in three of four young raccoons which had been naturally confined to chimneys. Two of these four young raccoons were infested with a nest-dwelling tick, Ixodes texanus.

An indirect immunofluorescence test was used during 1982-1983 to identify antibodies to Lyme disease spirochetes in humans, white-footed mice, and raccoons. Serologic tests detected IgM or total Ig antibodies in serum samples from 67 persons. Onset of illness, as marked by erythema chronicum migrans (ECM), occurred mainly during July and August. The majority of the persons with Lyme disease lived in south central and southeastern Connecticut. Analyses also verified prior spirochetal infections in 29 of 323 (9 percent) white-footed mice and in three of 34 (9 percent) raccoons captured at sites with or without evidence of human infections. Results indicate potential for Lyme disease at numerous localities in Connecticut.

Fundamental to maintaining a healthy environment in a mechanically ventilated livestock building is a continuous flow of air, with at least 4 exchanges per hour to remove moisture and aerosol contaminants. A correctly designed fresh air intake system is mandatory for uniform air distribution. Proper insulation is essential. Supplemental heat may be required to permit continuous ventilation while maintaining an ambient temperature suitable for preserving animal health.

Immature and adult ixodid ticks were collected during 1983 and 1984 in Newtown, Connecticut, an area endemic for Rocky Mountain spotted fever (RMSF), to determine prevalence of infection by spotted fever group (SFG) rickettsiae. Direct fluorescent-antibody (FA) staining revealed SFG organisms in 6 (1.8%) of 332 Dermacentor variabilis larvae, 5 (7.8%) of 64 D. variabilis nymphs, and in 2 (40%) of 5 Ixodes cookei nymphs removed from small- and medium-sized mammals. Hemolymph tests detected rickettsia-like organisms in 15 (8.8%) of 170 D. variabilis adults; 8 specimens retested by direct FA were negative. In contrast, hemocytes from 5 (8.6%) of 58 Ixodes texanus females contained organisms that stained positively in both hemolymph and direct FA tests. An indirect microimmunofluorescence test identified specific antibodies to Rickettsia rickettsii, the etiologic agent of RMSF, in serum samples from a chipmunk, raccoons, and white-footed mice. Results indicate that immature or adult ticks of at least three species may be involved in the maintenance and transmission of SFG rickettsiae at Newtown.

Tissue, external and whole animal insulation values were determined for 12 newborn male Holstein calves continuously housed for two weeks in hutches within environmental chambers in which temperature was maintained at a constant 17 degrees C (three calves) or cycled on a daily basis either between -20 degrees and -8 degrees C (three calves) or between -30 degrees and -18 degrees C (six calves). Three of the six calves at the coldest temperature were outfitted with an insulated coat. The insulated coat provided calves a 52% increase in total insulation. Tissue insulation of cold-housed calves increased 37.2% over the first two weeks of life. It was concluded that the capacity for vasoconstriction improved with age. External insulation did not change significantly except during the first week in cold-housed calves without insulated coats. External insulation values were five to eight times those of tissue insulation values for all treatment groups. This indicated that insulation of structures external to the skin (hair, bedding, ground, etc.) provided most of the insulation for calves.

A total of 612 Peromyscus leucopus, 11 Microtus pennsylvanicus, 21 Clethrionomys gapperi, and 4 Tamias striatus was collected in Connecticut and examined for Babesia and Grahamella during 1976 and 1977. Babesia antibodies were detected in sera of 9 P. leucopus collected from 4 sites. Babesia parasites were not detected in the blood smears of captured rodents. Subsequent splenectomy and subinoculation of blood from these rodents into susceptible animals failed to induce disease and no Babesia was isolated. Six of 10 P. leucopus inoculated with a Shelter Island, New York strain of B. microti remained infected for 3 1/2 months. Indirect fluorescent antibody titers of experimentally infected P. leucopus ranged from 1:8 to 1:256. Prevalence of Grahamella peromysci infection, as determined from examinations of blood smears of P. leucopus, was 13%. This infection rate is a conservative estimate because parasitemia is difficult to detect in intact animals. Twenty of 58 P. leucopus, taken at 2 sites and with negative blood smears for G. peromysci, developed parasitemia after splenectomy.

Spirochetes were isolated from 71 subadult Ixodes dentatus removed from cottontail rabbits captured in Millbrook, N.Y., and in New York, N.Y. Spirochetes were also cultured from kidney tissues of six rabbits. While all isolates reacted with monoclonal antibody H9724, which identifies the spirochetes as borreliae, more than half did not bind with antibody H5332 and even fewer reacted with H3TS, both of which were produced to outer surface protein A of Borrelia burgdorferi. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profiles of three isolates differed from one another and from all previously characterized B. burgdorferi strains from humans, ticks, and wildlife in North America. The 12 periplasmic flagella that originated subterminally from each pointed end of a rabbit Borellia isolate contrasted with the 11 or fewer flagella for B. burgdorferi reported previously from North America. Although DNA homology and restriction endonuclease analysis also revealed differences among a rabbit kidney isolate, an I. dentatus isolate, and B. burgdorferi B31, similarities were sufficient to lead us to conclude that the borreliae in rabbits and I. dentatus are B. burgdorferi. Enzyme-linked immunosorbent assay titers of sera from humans with diagnosed Lyme disease to rabbit tick B. burgdorferi were often similar to one another and to those recorded for a reference B. burgdorferi strain.

Histologic detection of micrometastatic melanoma in the placenta led to the clinical diagnosis in the mother. Metastases were present in the intervillous space and in villous stroma. The infant has survived and has no evidence of disease at one year of age. The mother died of metastatic disease 6 months post-partum. Literature review shows that only 25% of infants with placental metastatic melanoma succumb to the disease. In a review series of 16 cases, there are some maternal clinical factors which seem to have some bearing on the likelihood of fetal metastasis and death from disease. Features that are associated with an unfavorable fetal/infant prognosis are: maternal age less than 30 years, primiparity, leg primary site, disease onset greater than 3 years prior to pregnancy, node-1 metastatic status prior to pregnancy, M4 status in third trimester, birth at greater than 36 weeks' gestation, male sex. Extent of penetration of metastases into the villi does not have prognostic significance.

The energy content of spider eggs was determined on samples from 12 species representing 6 families. These values ranged from 26.32 to 29.00 joules per mg ash-free dry mass with a mean of 27.30±SE of 0.27. The higher values were found in those species that overwinter as developmental stages within the egg sac. Rates of energy expenditure of developing eggs and spiderlings within the egg sac were only 7 to 19% of those of emerged spiderlings. The energy conserved by this reduction in rate of metabolism may facilitate survival without feeding during the potentially long periods of aerial dispersal by ballooning, a characteristic activity of most newly emerged spiderlings. The variation in mass-specific energy content is less than variation in clutch size and individual egg size on an intra-and interspecific basis. There was no correlation between energy content per unit egg mass and size of the female parent, egg size, or clutch size. Further analysis indicated that no single measure such as clutch size accurately represents the proportional amount of energy invested in reproduction in these animals.

The relationships between dimensions of book lung subunits were measured and analyzed as a function of body size in diverse spiders over a body mass range of 3.4 to 3,190 mg. Book lungs are the characteristic respiratory gas exchange organs in these arachnids. Actual gas exchange occurs across numerous air-filled cuticular plates, which invaginate hemolymph sinuses within the abdomens of these animals. Characteristic linear dimensions of these air-filled compartments reflecting diffusion paths scaled to the 0.2 power of body mass and showed only a fourfold increase over the size range in the sample. This deviation from isometric scaling in the direction obtained and its numerical similarity to scaling of alveolar dimensions to body size in vertebrates was interpreted as an adaptation to reduce diffusion distances between these compartments and vascular fluids. Conversely, lengths and widths of these plates scaled to the one-third power of body mass, isometric scaling, and increased between six-and eightfold over the size range. This result is consistent with the hypothesis that respiratory gas distribution within spider lungs is achieved by convective mixing as has been recently hypothesized., (Copyright © 1980 Wiley-Liss, Inc.)

Three parameters were used in 1976 and 1977 to assess the status of Rocky Mountain spotted fever (RMSF) in Connecticut--compilation and review of clinical data on suspected human cases for the 13-year period 1965--1977, examination of tick tissues for spotted fever-group rickettsiae by the hemolymph test and direct immunofluorescence, and analyses of mammalian sera for antibodies against Rickettsia rickettsii. There were six presumptive RMSF cases which probably originated in Connecticut. Four of these cases occurred in areas where the American dog tick, Dermacentor variabilis, abounds. A total of 2994 ticks were examined by the hemolymph test. Rickettsia-like organisms were observed in 67 (2.9%) of 2330 D. variabilis and two (0.6%) of 351 Ixodes sp. near scapularis. Fewer than one-half of these organisms stained positively with spotted fever-group conjugate. Microagglutination tests on 1093 mammalian sera indicated that eight (16%) of 49 raccoons, 14 (2.6%) of 549 white-tailed deer, eight (1.7%) of 470 white-footed mice, and one of two gray squirrels had agglutinins in titers greater than or equal to 1:8 against R. rickettsii. Spotted fever-group rickettsiae are present at low frequency in inland as well as coastal regions of Connecticut.