The American Journal of Tropical Medicine and Hygiene - Volume 33, Issue 5, 1984

The migration of Schistosoma mansoni cercariae and schistosomules, and the cellular responses of the hosts to these parasites, in primary and challenge infections, were examined and cine-recorded in vivo in the hypodermal tissues in Algire chambers which had been inserted in dorsal skins of C3H/HeJ mice and cheek pouches of Syrian hamsters, and checked in serial sections. In hamsters, many cercariae penetrated blood vessels within minutes after being injected into their chambers, in contrast to mice, in which penetration into vessels was scanty during the first hour. Schistosomule penetration into blood vessels was slow in both species. To ascertain whether or not the cercariae that had rapidly penetrated blood vessels in hamsters were capable of producing the disease, the chambers were removed a few hours after infection had been induced. Eight to 10 weeks later it was found that the parasites had produced the disease, since egg granulomas were found in the liver of hamsters. The cellular reactions of the hosts to the parasites revealed that granulocytes adhered first and most extensively to the tails of cercariae in essentially equal amounts in primary and challenge infections, but the adherence of such cells to the body of cercariae was geater in challenge infections. Nevertheless, an appreciable number of cercariae in both naive and challenge-infected animals were free of granulocytes or had less than five cells adhering to them. Furthermore, granulocyte adherence to cercariae or schistosomules was not necessarily permanent; the latter were usually free of cells a few hours after infection had been induced. Another difference between naive and challenge infections was that cercarial motility was more rapidly depressed in the latter infections.

Schistosomula and adults of Schistosoma mansoni were grown from cercariae in cultures differing only in the treatment of the red blood cells fed to the organisms. “Pink ghosts,” containing about 5% of the original hemoglobin, were produced by hemolysis in water; “white ghosts” with no detectable hemoglobin were made in 5 mM phosphate buffer, pH 8. Early growth and development were more rapid and vigorous, and pairs formed more readily when pink ghosts, rather than intact erythrocytes were fed. Schistosomula remained stunted and undeveloped when fed with white ghosts. Attempts at reconstitution of the latter by addition of hemoglobin, concentrated erythrocyte lysate, or pressure-liquefied pink ghosts did not restore growth-promoting activity. Pink ghost-fed worms, particularly paired males, attached to the dish bottom by their acetabulum and oral sucker and travelled by an active looping motion. Substrates of collagen or fibrin or a mammalian cell monolayer did not affect this behavior. Such attachment and locomotion are interpreted as instinctive migratory behavior of schistosomes.

Six Schistosoma mansoni isolates obtained from the same endemic area were compared experimentally; three were from patients with the hepatosplenic form and three from patients with the intestinal form of schistosomiasis. The following features were analyzed in mice: body weight, liver and spleen weights, mortality, infectivity, number of S. mansoni eggs/female worm, egg distribution in the viscera, and hemogram. Three groups of mice, used as controls, were infected with the LE strain of S. mansoni which is routinely maintained in the laboratory. The group of isolates from patients with the hepatosplenic form of schistosomiasis did not show important differences in relation to those from non-hepatosplenic patients; in almost all features analyzed the differences among these six isolates were within the variation limits observed with the LE strain of S. mansoni.

Through preliminary experiments, an effective method for radiolabeling Brugia malayi-infected mosquitoes in order to produce labeled infective Brugia larvae was developed. Starting on the 6th day after the infective blood meal, mosquitoes were fed a 7% sucrose solution containing 100 µCi/ml 75Se-L-methionine for 5 days. Infective larvae, retrieved 2 days after this labeling period, averaged 381 ± 136 counts/min. Jirds were infected with these infective, labeled larvae either by allowing infected mosquitoes to feed on uninfected jirds for 30 min or by inoculating jirds subcutaneously in the groin with washed larvae recovered from mosquitoes. Jirds were killed at various times after infection and were sliced into approximately 0.5 mm thick sagittal sections, which were dried and placed on X-ray film. Autoradiograms were developed after 30–60 days at 5°C. In a sample of 26 inoculated jirds, approximately 30% of the infecting larvae could subsequently be accounted for as Ag° foci on autoradiograms. The Ag° foci representing larvae were apparent up to 2.5 weeks after infection. In jirds infected by mosquito feeding, the Ag° associated with the feeding site persisted for more than 6 weeks after infection.

This report describes a case of tropical pyomyositis in a 25-year-old man in which gray scale ultrasound played a critical role in the diagnosis and management. A review of relevant literature is provided, and a recommended approach to the diagnosis of this disease is given.

To determine the modes of transmission of cholera in the regency of Pidie, Indonesia, and to consider strategies for its control, we set up a laboratory to identify Vibrio cholerae 01 from patients with severe diarrhea in all government clinics in the regency and questioned culture-positive cases and neighborhood controls about possible exposures to V. cholerae 01. Between 12 July and 15 August 1982, 63 of 138 suspected cholera cases were confirmed by the laboratory; 53 of these patients were seen and followed up. We were unable to identify a single, indisputable mode of transmission for cholera which was amenable to immediate control. Nonetheless, a number of factors, including exposure to water from the Tiro-Sigli River and consumption of ice, were associated with disease. Other findings bring into question the value of current practices of chlorinating dugwells and disinfecting homes with Lysol® during a cholera outbreak. The case-control approach to investigating the mode of transmision of cholera has distinct limitations when applied in endemic setting where there may not be a single predominant vehicle of transmission, or where the vehicle such as river water is used by all and is only periodically contaminated.

Serum samples were obtained within 3 days of capture from 106 cynomolgus monkeys (Macaca fascicularis) in peninsular Malaysia. Fifty-two monkeys were trapped on the fringes of palm oil estates and 54 in dense primary jungle. Sera were tested for antibodies to hepatitis A virus (HAV) with a commercial radioimmunoassay. Twenty-four animals had detectable serum anti-HAV activity (6 of 52 from palm oil estate sites and 18 of 54 from primary jungle sites). Among monkeys at both sites, antibody prevalence was strongly correlated with animal weight: overall only four of 69 monkeys (6%) weighing less than 2.0 kg had serum anti-HAV antibodies, while 14 of 29 (48%) weighting 2.0 to 3.9 kg, and 6 of 8 (75%) weighing 4.0 kg or more, had serum anti-HAV antibodies. These data suggest that wild cynomolgus monkeys in Malaysian jungles become infected with HAV or an HAV-like virus at a rate comparable to that of humans in the same region, and raise the possibility of a sylvatic cycle for HAV.

A cell culture-adapted variant of the rabies-related Mokola virus was demonstrated to replicate in inoculated Aedes aegypti mosquitoes. Replication was slow compared to many arboviruses in their vectors. Maximum titers were not obtained until after approximately 6 weeks of extrinsic incubation. Mokola virus underwent nine mosquito-mosquito passages at approximately monthly intervals and was thus maintained in insects for 340 days before terminating the study. Virus antigen was detected by immunofluorescence in a variety of mosquito tissues and organs, including salivary glands, but primarily in nervous tissue. Irrefutable virus transmission by bite could not be demonstrated because of equivocal results. Transovarial passage of virus was observed in the mosquito. Viremia in baby mice was demonstrable. Ornithodoros moubata nymphal ticks were exposed to viremic mice but failed to become infected.

Barriers to dissemination of Middle American epizootic hemagglutination inhibition subtype I-AB, and enzootic, subtype IE, Venezuelan encephalitis (VE) viruses were examined in a colony of the enzootic vector mosquito, Culex (Melanoconion) taeniopus. This species is highly susceptible to oral infection with enzootic, but not epizootic, virus strains. Adult female mosquitoes were intrathoracically inoculated with epizootic virus suspensions to ascertain whether a mesenteron infection barrier exists to these subtype I-AB strains. All inoculated mosquitoes became infected, including those receiving only 10 chick embryo cell culture plaque-forming units (CEC pfu). This confirmed that a mesenteron infection barrier exists to epizootic, but not enzootic Middle American VE strains. Mosquitoes were also given high titer hamster bloodmeals of epizootic viruses and dissected at 2-day intervals to determine the location of virus in the few infected individuals. With mean bloodmeal titers of up to 105.3 CEC pfu, only 20% or less of the mosquitoes became infected, and virus replication was confined to the mesenteron. This indicated that a mesenteron escape barrier to epizootic VE viruses exists in this mosquito. Mosquitoes were also given large and small oral doses of enzootic virus strains to compare viral replication patterns. With high titer bloodmeals, virus disseminated from the mesenteron within 4 days of infection, and titers in mosquitoes peaked 7–9 days after infection. All mosquitoes that ingested large doses became infected. Mosquitoes receiving small oral doses of enzootic viruses showed a different pattern of virus replication. Over 90% of mosquitoes engorging bloodmeal titers of 100.7 CEC pfu became infected, but the virus failed to escape from, or was delayed in disseminating beyond, the mesenteron. Thus, the mesenteron escape barrier also exists to enzootic strains, and appears to be inversely related to the bloodmeal titer, reaching 61% with undetectable bloodmeal titers.

Transmission electron microscopy was used to determine if eastern equine encephalitis (EEE) virus infects and replicates in the salivary glands of Culiseta melanura after 3 days of extrinsic incubation (EI). The Cs. melanura studied were from a colony strain, were orally infected, and had EI periods of 55–69 hours. Both naked nucleocapsids and enveloped virions were present in aggregates, suggestive of viral replication, within salivary gland acinar cells. Nucleocapsids were present in the cytoplasm below the plasma membrane that lined apical cavities. Enveloped virions occurred in the salivary matrix within apical cavities. Some nucleocapsids appeared to be budding through the plasma membrane around apical cavities and maturing into infectious virions. These results suggest that Cs. melanura is capable of biological transmission of EEE virus after ≤ 3 days of EI.

Enzyme immunoassays (EIAs) producing either chromogenic or fluorogenic end products were developed and evaluated for detection of eastern equine encephalomyelitis (EEE) and Highlands J (HJ) viruses in pools of Aedes triseriatus mosquitoes. Overnight incubation of the mosquito samples in the EIA significantly enhanced the sensitivity of the test. Both the EEE and HJ EIAs were sensitive, readily detecting one infected mosquito in a pool with 99 noninfected, and specific, distinguishing homologous from the alternate alphavirus and other arboviruses. By 3 days post-infection after intrathoracic inoculation, EEE virus was isolated from 100% (30/30) of the mosquitoes examined. Concurrently, EEE virus antigen was detectable by EIA in 100% (30/30) of examined mosquitoes and by indirect fluorescent antibody (IFA) technique in 77% (23/30) of the examined mosquito head squash preparations.

Enzyme immunoassays (EIAs) for eastern equine encephalomyelitis (EEE) and Highlands J (HJ) virus antigens were compared in a retrospective study with standard virus isolation procedures (VIP) for detection of alphavirus-infected mosquito pools. The original VIP was a plaque assay in chick embryo cell culture, and was performed in the years from 1979 to 1981. Using the original VIP as the reference standard, the sensitivity rate of the EIA was 0.7674 and the false negative rate was 0.2326. However, when the storage age and the initial virus titer of the sample were considered, the sensitivity rate increased. For samples containing > 1,500 plaque-forming units (PFU) per ml of virus during the original VIP, the sensitivity rate of the EIA was 0.97; but the rate declined to 0.14 for those originally containing < 500 PFU per ml. Most of the false negatives (68%) occurred with samples containing < 500 PFU per ml. Presumably the low quantitites of virus in these 50 pools were lost during storage and handling; virus was obtained from only 16% (8/50) during reisolation attempts using BHK-21 cells. Specificity of the EIA was excellent; no false positive results were obtained and serological identification was identical to that determined by plaque reduction neutralization in > 98% of the pools examined. Characteristics of the pools, such as pool size, species of mosquitoes, or gravidity did not affect the EIA results. These studies support the use of EIAs in surveillance programs attempting to determine infection rates of known arboviruses in vector populations.

We studied the transmission of Fort Morgan (FM) virus within colonies of nesting Cliff Swallows and House Sparrows under three bridges in Morgan County, Colorado during 1976. Nests were examined, and blood or brain specimens were collected from nestlings once or twice a week. Flying birds and small mammals were also studied. We analyzed nesting activity, virus isolations from nestlings of both species, fledging success, multiple infections within a brood of nestlings, infection frequency by age of nestlings, nestling mortality, and infection frequencies by avian species and bridge site. Fort Morgan virus was isolated from 7% (80/1, 156) of the blood and brain samples collected from nestlings. The duration of viremia for nestling House Sparrows was at least 3–4 days based on virus isolation from sequential blood samples. Viremia of nestling Swallows and House Sparrows did not reduce fledging success, nor were young nestling sparrows viremic more frequently than older nestling sparrows. Nest destruction (by falling down) was a more important cause of nestling mortality than FM virus infection. All age groups of nestling sparrows were viremic at equal rates, but younger nestlings (≤7 days old) were more likely than older nestlings (> 7 days old) to develop an encephalitic infection. Among nestling House Sparrows, FM virus infections were clustered in time and space. Nestling House Sparrows with FM virus-infected nest-mates were infected more often than conspecifics whose nest-mates were not infected. We concluded that nestling Cliff Swallows and symbiotic House Sparrows that reside in swallow nesting colonies are the principal vertebrate hosts for the maintenance and amplification of FM virus.

Recent observations that Rattus rodents in Asia harbor Hantaan (HTN)-like viruses associated with hemorrhagic fever with renal syndrome led us to investigate the possibility that similar viruses might be present in the United States. Wharf rats were captured at major port cities, their sera were examined for HTN antibody by immunofluorescent antibody (IFA) assay and tissues from selected antibody-positive rats were examined for HTN antigen. Rats positive for both antibody and antigen to HTN virus were found in Philadelphia, PA and Houston, TX. Infected rats were found clustered in discrete foci where a significant proportion was antibody and antigen positive. Viruses isolated from lung tissues of Rattus norvegicus captured at Philadelphia and Houston were grown in cell culture and shown to be closely related to each other and to the 76–118 strain of HTN virus by IFA, but clearly distinct from HTN by plaque reduction neutralization tests. The isolates appear to be variants of the same new virus, for which the name Girard Point virus is proposed.

Two new rhabdoviruses, designated Carajas and Maraba, are described. Both were isolated from phlebotomine sand flies (Lutzomyia spp.) collected in the Amazon basin of Brazil. One recovery of Carajas virus was made from male sand flies. By complement-fixation and neutralization tests both agents were shown to be members of the vesicular stomatitis virus (VSV) serogroup (genus Vesiculovirus). The pathogenicity of the two viruses in mice and Vero cells is similar to that of VSV-Indiana and VSV-New Jersey. Both Carajas and Maraba viruses replicated in Lutzomyia longipalpis following intrathoracic inoculation, and both agents were transovarially transmitted in this sand fly species.

This paper describes a series of experiments which were done to determine the behavior of 14 different phleboviruses in laboratory-reared sand flies (Phlebotomus papatasi, P. perniciosus and Lutzomyia longipalpis) after oral and parenteral infection. Most of the viruses replicated in the sand flies after intrathoracic inoculation; however, the insects were quite refractory to oral infection. Six of 11 phleboviruses tested were transovarially transmitted in one or more sand fly species. The percentage of infected F1 offspring produced by parenterally infected female parents ranged from 1.5–60%, depending on the virus type used. These data support the hypothesis that some of the phleboviruses are maintained in sand flies by transovarial transmission.

Quantitative studies of selected features of peripherally induced Rickettsia mooseri (=R. typhi) infection in Rattus norvegicus-derived white laboratory rats revealed a unique association between microbe and amplifying vertebrate host which appears to be especially conducive to maintenance of the enzootic cycle. Both adult and newborn (1–3 days old) rats were highly susceptible to percutaneous infection (ID50 = ∼1 organism), but neither showed signs of disease or died even when inoculated with 104–105 plaque-forming units. Gain in body weight of infected newborn rats was indistinguishable from that of uninfected newborn rats over the first 3 weeks of life. The course of the systemic infection, as measured by the rise and fall of R. mooseri titers in blood, brain and kidney and the serum antibody response, was almost identical in adult and newborn rats. Thus, despite their immaturity in certain immunological processes, newborn rats controlled postnatal R. mooseri infection about as well as did adult rats. The rickettsemic period of about 10 days corresponds to the period of infectivity of inoculated rats for fleas. Rickettsiae were not isolated from blood, brain or kidneys by methods employed for more than 4–5 weeks after infection. Serum antirickettsial antibodies persisted for at least 60 weeks postinfection, i.e., longer than the usual life span of rats in nature and, hence, are a valid measure of the cumulative experience of rat populations with R. mooseri infection.

A survey for the prevalence of Rocky Mountain spotted fever (RMSF) antibodies in dogs associated with confirmed human cases in Ohio was conducted during 1981. Twelve of 14 confirmed cases (85%) had a history of dog association prior to onset of RMSF. A total of 29 dogs were included in the study, with 16 dogs providing serum samples for antibody testing and the remainder providing tick samples. Serum samples tested by indirect microimmunofluorescence techniques revealed 12/16 dogs (75%) to be seropositive for Rickettsia rickettsii. A total of 310 ticks were collected from study dogs and the vegetation surrounding RMSF case exposure sites. Twenty-two (7.1%) of these ticks (all Dermacentor variabilis) were found to be infected with spotted fever group rickettsiae. Four ticks (1.3%) were infected with R. rickettsii, 13 (4.2%) with Rickettsia montana, and four (1.3%) with Rickettsia bellii. R. montana, a nonpathogen, was the only rickettsia found in dogs (antibodies) and ticks (isolation) associated with human cases in Southern Ohio.

A nematode in a subcutaneous nodule from the anterior chest of a 12-year-old boy in Northern Thailand was identified as a third-stage larval dioctophymatid, possibly Dioctophyma renale, the second such larva to be reported from man.

Spider poisoning is usually a result of the direct bite, envenomation through the chelicera, of the spider. Reported is an unusual case in which poisoning occurred through the conjunctiva when a fragment from a smashed black widow spider (Latrodectus hesperus) flew into a patient's eye. In addition to an immediate local reaction involving the periorbital tissues, the patient experienced systemic effects.