A spatio-temporal analysis of scrub typhus and murine typhus in Laos; implications from changing landscapes and climate.

BACKGROUND: Scrub typhus (ST) and murine typhus (MT) are common but poorly understood causes of fever in Laos. We examined the spatial and temporal distribution of ST and MT, with the intent of informing interventions to prevent and control both diseases. METHODOLOGY AND PRINCIPLE FINDINGS: This study included samples submitted from 2003 to 2017 to Mahosot Hospital, Vientiane, for ST and MT investigation. Serum samples were tested using IgM rapid diagnostic tests. Patient demographic data along with meteorological and environmental data from Laos were analysed. Approximately 17% of patients were positive for either ST (1,337/8,150 patients tested) or MT (1,283/7,552 patients tested). While both diseases occurred in inhabitants from Vientiane Capital, from the univariable analysis MT was positively and ST negatively associated with residence in Vientiane Capital. ST was highly seasonal, with cases two times more likely to occur during the wet season months of July-September compared to the dry season whilst MT peaked in the dry season. Multivariable regression analysis linked ST incidence to fluctuations in relative humidity whereas MT was linked to variation in temperature. Patients with ST infection were more likely to come from villages with higher levels of surface flooding and vegetation in the 16 days leading up to diagnosis. CONCLUSIONS: The data suggest that as cities expand, high risk areas for MT will also expand. With global heating and risks of attendant higher precipitation, these data suggest that the incidence and spatial distribution of both MT and ST will increase.

CD4+ T Cells Are as Protective as CD8+ T Cells against Rickettsia typhi Infection by Activating Macrophage Bactericidal Activity.

Rickettsia typhi is an intracellular bacterium that causes endemic typhus, a febrile disease that can be fatal due to complications including pneumonia, hepatitis and meningoencephalitis, the latter being a regular outcome in T and B cell-deficient C57BL/6 RAG1-/- mice upon Rickettsia typhi infection. Here, we show that CD4+ TH1 cells that are generated in C57BL/6 mice upon R. typhi infection are as protective as cytotoxic CD8+ T cells. CD4+- as well as CD8+-deficient C57BL/6 survived the infection without showing symptoms of disease at any point in time. Moreover, adoptively transferred CD8+ and CD4+ immune T cells entered the CNS of C57BL/6 RAG1-/- mice with advanced infection and both eradicated the bacteria. However, immune CD4+ T cells protected only approximately 60% of the animals from death. They induced the expression of iNOS in infiltrating macrophages as well as in resident microglia in the CNS which can contribute to bacterial killing but also accelerate pathology. In vitro immune CD4+ T cells inhibited bacterial growth in infected macrophages which was in part mediated by the release of IFNγ. Collectively, our data demonstrate that CD4+ T cells are as protective as CD8+ T cells against R. typhi, provided that CD4+ TH1 effector cells are present in time to support bactericidal activity of phagocytes via the release of IFNγ and other factors. With regard to vaccination against TG Rickettsiae, our findings suggest that the induction of CD4+ TH1 effector cells is sufficient for protection.

Opossums and Cat Fleas: New Insights in the Ecology of Murine Typhus in Galveston, Texas.

Murine typhus is an acute undifferentiated febrile illness caused by Rickettsia typhi The classic reservoir (Rattus spp.) and flea vector (Xenopsylla cheopis) were once culprits of murine typhus in the United States. Vector and rodent control efforts have drastically decreased the prevalence of disease, except in a few endemic foci where opossums and cat fleas play a role in transmission. Since 2012, there has been a reemergence of murine typhus in Galveston, TX. We hypothesize that opossums and cat fleas are involved in the transmission of R. typhi in Galveston. To explore this, we sought to find the seroprevalence of typhus group antibodies from opossums. We also sought to find the prevalence of R. typhi in fleas parasitizing these animals. We collected blood from 12 opossums and found that eight (66.7%) had the presence of anti-R. typhi antibodies. All opossums were infested with fleas; a total of 250 Ctenocephalides felis fleas were collected from these animals. Seven opossums (53.8%) were infested with fleas that had molecular evidence of R. typhi infection, while six (46.2%) were infested with fleas that contained Candidatus Rickettsia senegalensis, an organism closely related to R. felis The minimum flea infection rate for R. typhi was 7.0%. The minimum infection rate for Candidatus R. senegalensis was 6.1%. Our study demonstrates that fleas infected with R. typhi parasitize opossums in Galveston. It is therefore likely that opossums and their fleas play a role in the city's recent reemergence of murine typhus.

The role of dogs in the eco-epidemiology of Rickettsia typhi, etiological agent of Murine typhus.

Rickettsia typhi, etiological agent of Murine typhus (MT), is transmitted to humans from an animal reservoir through two cycles: a classic rat-flea-rat cycle, and a peridomestic animal cycle. There are not many studies concerning which animals are involved in the peridomestic cycle, and most of them are focused on cats. The aim of this study was to determine the presence of R. typhi in dogs, not only by serological methods but also by direct methods such as culture and molecular detection. Two hundred and one dog blood samples were collected from Veterinary clinics, kennels, and shelters in Northeastern Spain (2006-2008). Age, sex, municipality, living place, healthy status, contact with animals, and ectoparasite infestations were surveyed. IgG was measured by IFA. Titers ≥ 1/64 were considered positive. Cultures were carried out using samples of dogs with titers ≥ 1/128. The molecular detection was performed by real-time PCR. Nine dogs (4.5%) were positive according to IFA (5: 1/64; 3: 1/128; 1: 1/512). There were no significant differences in the rates of antibodies related to any of the variables. Rickettsial DNA was detected in two cultures. Sequences obtained were identical to those of R. typhi. The results show direct and indirect evidences of the presence of R. typhi infection in dogs.

Transmission of flea-borne zoonotic agents.

Flea-borne zoonoses such as plague (Yersinia pestis) and murine typhus (Rickettsia typhi) caused significant numbers of human cases in the past and remain a public health concern. Other flea-borne human pathogens have emerged recently (e.g., Bartonella henselae, Rickettsia felis), and their mechanisms of transmission and impact on human health are not fully understood. Our review focuses on the ecology and epidemiology of the flea-borne bacterial zoonoses mentioned above with an emphasis on recent advancements in our understanding of how these organisms are transmitted by fleas, maintained in zoonotic cycles, and transmitted to humans. Emphasis is given to plague because of the considerable number of studies generated during the first decade of the twenty-first century that arose, in part, because of renewed interest in potential agents of bioterrorism, including Y. pestis.

Murine typhus returns to New South Wales: a case of isolated meningoencephalitis with raised intracranial pressure.

Murine typhus (MT) occurs worldwide, but, in Australia, is only regularly diagnosed in south-west Western Australia. Meningoencephalitis is an uncommon complication of MT, often accompanied by rash or systemic involvement. We report a case of MT presenting exclusively with meningoencephalitis, raised intracranial pressure, papilloedema and bilateral 6th cranial nerve palsies. MT should be considered in patients with "aseptic" meningitis or meningoencephalitis, even in the absence of other typical features of a typhus-like illness.

[Experimental study of the inoculative transmission of Rickettsia typhi by gamasid mites (Gamasidae) Ornithonyssus bacoti].

The authors' studies have established that the concentration of Rickettsia typhi may increase about 100-fold in the infected Ornithonyssus bacoti mites. At the time, when on feeding 20 to 200 adult mites on guinea-pigs and albino rats 4 to 36 days after inoculation, they did not transmit Rickettsia typhi on blood sucking.

[The history of the flea in art and literature]. / La storia della pulce nell'arte e nella letteratura.

The flea has been, indirectly, one of the protagonists in the history of man. As one of the two vectors of Yersinia pestis, the etiological agents of the Black Death, the flea (Xenopsylla cheopis) has contributed, over the centuries, to the death of millions of people in many countries. Galileo Galilei was the first to observe the flea with a microscope (1624), but the credit of depicting it with a stunning drawing goes to the Britisher Robert Hooke in 1665. A number of zoologists, including Antonie Van Leeuwenhoek and Diacinto Cestoni, well described and illustrated the life cycle of the flea in the XVII century. Some of these reports inspired scholars such as J. Swift and J. Donne for the composition of classic poems. Also, the flea, alone and with its hosts, has inspired a number of artists to create fine paintings; among them: G. M. Crespi, G. B. Piazzetta, G. de la Tour and others. Colorful sonnets on the flea in the Roman dialect were written by G. Belli and Trilussa. The flea also, as a theme, inspired musicians such as G. F. Ghedini and M. Mussorgsky, play writers such as Feydeau and moviemakers such as Charlie Chaplin. The flea is, indissolubly, connected with the history of Black Death. This disease in man is, in fact, caused--as demonstrated by Yersin and Simond--by the triad: bacterium (Yersinia pestis)/rat/flea (Xenopsylla cheopis). Over the centuries, Black Death has had a deep impact on both the visual arts and literature and, as a result, a very large number of paintings and other works of art have been produced to remember these tragic episodes. In the field of literature, Black Death has been skillfully described by writers such as Boccaccio, Manzoni and Camus. Finally, in recent years, following the discovery of the existence of a large market for the control of fleas in small animals, the interest in this minute insect has been resurrected and, parallel to that, the rebirth of the flea iconography, through electromicroscopy, has also taken place.

Rickettsial actin-based motility: behavior and involvement of cytoskeletal regulators.

Actin-based motility (ABM) is employed by spotted fever group (SFG) rickettsiae, such as Rickettsia rickettsii, to promote cell-to-cell spread. Time-lapse video microscopy revealed that ABM is not strictly confined to SFG rickettsiae as typhus group R. typhi moved at approximately the same rate as R. rickettsii (approximately 4 micro m/min), but in a highly erratic fashion. A number of common behaviors were observed between ABM of R. typhi and R. rickettsii, such as entrance into plasma membrane protrusions, formation of new actin tails only on the old surface of newly formed daughter cells, and quick (within 15 sec) reassembly of the actin tail to the opposite pole upon contact with cellular structures that impede forward movement. This last behavior suggests that the rickettsial protein(s) required for ABM is uniformly localized to both poles of the bacterium and possibly throughout the rickettsial surface. Functional roles in rickettsial ABM for neuronal Wiskott-Aldrich syndrome protein (N-WASP) and the actin-related protein (Arp)2/3 complex, critical regulators of ABM of other pathogens, have not been established. Domains of N-WASP that have characterized inhibitory effects on N-WASP or Arp2/3 complex function were expressed in HeLa cells infected with R. rickettsii. Shigella flexneri-infected cells were used as a control. When ectopically expressed, the VCA domain of N-WASP (VCA) acts as a dominant/negative with respect to Arp2/3 complex function and N-WASP missing VCA (DeltaVCA) acts as a dominant/negative form of N-WASP. Expression of VCA or DeltaVCA severely inhibited S. flexneri ABM (no Shigella motility observed in the majority of expressing cells) while only moderately inhibiting ABM of R. rickettsii (approximately 35% decrease in the rate of ABM). In addition, ectopically expressed full-length GFP-N-WASP was recruited by S. flexneri but not R. rickettsii, and Arp3 was detected by indirect immunofluorescence in S. flexneri actin tails but not within R. rickettsii actin tails. Collectively, these data suggest that rickettsial ABM is independent of N-WASP and Arp2/3 complex function.

Rickettsia-macrophage interactions: host cell responses to Rickettsia akari and Rickettsia typhi.

The existence of intracellular rickettsiae requires entry, survival, and replication in the eukaryotic host cells and exit to initiate new infection. While endothelial cells are the preferred target cells for most pathogenic rickettsiae, infection of monocytes/macrophages may also contribute to the establishment of rickettsial infection and resulting pathogenesis. We initiated studies to characterize macrophage-Rickettsia akari and -Rickettsia typhi interactions and to determine how rickettsiae survive within phagocytic cells. Flow cytometry, microscopic analysis, and LDH release demonstrated that R. akari and R. typhi caused negligible cytotoxicity in mouse peritoneal macrophages as well as in macrophage-like cell line, P388D1. Host cells responded to rickettsial infection with increased secretion of proinflammatory cytokines such as interleukin-1beta (IL-1beta) and IL-6. Furthermore, macrophage infection with R. akari and R. typhi resulted in differential synthesis and expression of IL-beta and IL-6, which may correlate with the existence of biological differences among these two closely related bacteria. In contrast, levels of gamma interferon (IFN-gamma), IL-10, and IL-12 in supernatants of infected P388D1 cells and mouse peritoneal macrophages did not change significantly during the course of infection and remained below the enzyme-linked immunosorbent assay cytokine detection limits. In addition, differential expression of cytokines was observed between R. akari- and R. typhi-infected macrophages, which may correlate with the biological differences among these closely related bacteria.

Experimental transmission of murine typhus by Xenopsylla cheopis flea bites.

Transmission of Rickettsia typhi to rats by the bites of Xenopsylla cheopis (Rothschild) fleas was investigated. Procedures rigorously excluded the possibility of contamination of the host skin by flea faeces. Fleas with R. typhi infection (21-25 days post-infection) which fed through bolting cloth (45 min exposure to ten fleas) transmitted rickettsiae with a success rate of 20%. Infective fleas allowed free access to their host for 8 h (10-15 fleas/rat) gave transmission rates of 45-68%. They were also capable of inoculating R. typhi through a membrane of rat skin on a feeder. Only fleas which had been infected for 21 days or longer transmitted R. typhi orally. Oral transmission appeared to be the result of regurgitation of rickettsiae present in the foregut lumen rather than through salivary secretions.

Transmission of murine typhus rickettsiae by Xenopsylla cheopis, with notes on experimental infection and effects of temperature.

In studies on experimental infection of Rickettsia mooseri (= R. typhi) in Xenopsylla cheopis and laboratory rats, it was found that 10 days after the infectious feeding, the fleas were voiding feces that were infective to rats upon inoculation. The feces remained infective for at least the duration of the experiment, and a quantity as small as 0.2 micrograms of feces would result in seroconversion of 67% of the rats upon inoculation. Fleas were capable of transmitting the infection to rats as early as seven days after feeding on rickettsemic rats, but the rate of transmission was much higher late in the course of rickettsial development in the flea, e.g., virtually 100% by day 17. Fleas transmitted R. mooseri infection even when they fed on the host for a maximum of 30 min and were removed from the rats at least 25 min before they could be expected to deposit any feces. These and other data suggest that R. mooseri may be transmitted by X. cheopis by the feeding process, and not merely through contact with infective feces or crushed fleas. The ambient temperature had a profound effect upon rickettsial growth in the fleas. At 18 degrees C, the rickettsial content of the fleas was below detectable levels for at least ten days and remained low throughout, whereas at 24 degrees C and 30 degrees C the rickettsial titer was consistently two or three times greater. However, if, after six days, the fleas were transferred from an environment of 18 degrees C to one at 24 degrees C or 30 degrees C, the rickettsial growth increased by two or three logs within one week.