Neoehrlichia mikurensis s an emerging and vector-borne zoonosis: The first human disease cases were reported in 2010. Limited information is available about the prevalence and distribution of Neoehrlichia mikurensis in Europe, its natural life cycle and reservoir hosts. An Ehrlichia-like schotti variant has been described in questing Ixodes ricinus ticks, which could be identical to Neoehrlichia mikurensis.
Three genetic markers, 16S rDNA, gltA and GroEL, of Ehrlichia schotti-positive tick lysates were amplified, sequenced and compared to sequences from Neoehrlichia mikurensis. Based on these DNA sequences, a multiplex real-time PCR was developed to specifically detect Neoehrlichia mikurensis in combination with Anaplasma phagocytophilum in tick lysates. Various tick species from different life-stages, particularly Ixodes ricinus nymphs, were collected from the vegetation or wildlife. Tick lysates and DNA derived from organs of wild rodents were tested by PCR-based methods for the presence of Neoehrlichia mikurensis. Prevalence of Neoehrlichia mikurensis was calculated together with confidence intervals using Fisher's exact test.
The three genetic markers of Ehrlichia schotti-positive field isolates were similar or identical to Neoehrlichia mikurensis. Neoehrlichia mikurensis was found to be ubiquitously spread in the Netherlands and Belgium, but was not detected in the 401 tick samples from the UK. Neoehrlichia mikurensis was found in nymphs and adult Ixodes ricinus ticks, but neither in their larvae, nor in any other tick species tested. Neoehrlichia mikurensis was detected in diverse organs of some rodent species. Engorging ticks from red deer, European mouflon, wild boar and sheep were found positive for Neoehrlichia mikurensis.
Ehrlichia schotti is similar, if not identical, to Neoehrlichia mikurensis. Neoehrlichia mikurensis is present in questing Ixodes ricinus ticks throughout the Netherlands and Belgium. We propose that Ixodes ricinus can transstadially, but not transovarially, transmit this microorganism, and that different rodent species may act as reservoir hosts. These data further imply that wildlife and humans are frequently exposed to Neoehrlichia mikurensis-infected ticks through tick bites. Future studies should aim to investigate to what extent Neoehrlichia mikurensis poses a risk to public health.
Vector-borne disease; Emerging zoonoses; Candidatus N. mikurensis; I. ricinus; Anaplasma phagocytophylum
Neoehrlichiosis caused by “Candidatus Neoehrlichia mikurensis” is an emerging zoonotic disease. In total, six patients have been described in Europe, with the first case detected in 2007. In addition, seven patients from China were described in a report published in October 2012. In 2009, we diagnosed the first human case of “Ca. Neoehrlichia mikurensis” infection in the Zurich area (Switzerland). Here, we report two additional human cases from the same region, which were identified by broad-range 16S rRNA gene PCR. Both patients were immunocompromised and presented with similar clinical syndromes, including fever, malaise, and weight loss. A diagnostic multiplex real-time PCR was developed for specific detection of “Ca. Neoehrlichia mikurensis” infections. The assay is based on the signature sequence of a 280-bp fragment of the “Ca. Neoehrlichia mikurensis” 16S rRNA gene and incorporates a “Ca. Neoehrlichia mikurensis” species, a “Ca. Neoehrlichia” genus, and an Anaplasmataceae family probe for simultaneous screening. The analytical sensitivity was determined to be below five copies of the “Ca. Neoehrlichia mikurensis” 16S rRNA gene. Our results show that the assay is suitable for the direct detection of “Ca. Neoehrlichia mikurensis” DNA in clinical samples from, for example, blood and bone marrow. In addition, it allows for monitoring treatment response during antibiotic therapy. Using the same assay, DNA extracts from 1,916 ticks collected in four forests in close proximity to the patients' residences (<3 km) were screened. At all sampling sites, the minimal prevalence of “Ca. Neoehrlichia mikurensis” was between 3.5 to 8% in pools of either nymphs, males, or females, showing a strong geographic association between the three patients and the assumed vector.
‘Candidatus Neoehrlichia mikurensis’ is a tick-borne bacteria implicated in human health. To date, ‘Ca. Neoehrlichia mikurensis’ has been described in different countries from Africa, Asia and Europe, but never in Spain. However, according to the epidemiological features of the main vector in Europe, Ixodes ricinus, its circulation in our country was suspected.
A total of 200 I. ricinus ticks collected in the North of Spain were analyzed. DNAs were extracted and used as templates for PCRs targeting fragment genes for Anaplasma/Ehrlichia detection. The amplified products were sequenced and analyzed.
‘Ca. Neoehrlichia mikurensis’ was amplified in two specimens. Furthermore, Anaplasma phagocytophilum was detected in 61 samples analyzed.
The detection of ‘Ca. Neoehrlichia mikurensis’ in I. ricinus ticks from Spain indicates its circulation and the potential risk of contracting a human infection in this country.
‘Candidatus Neoehrlichia mikurensis’; Anaplasma phagocytophilum; Ixodes ricinus; Spain
“Candidatus Neoehrlichia mikurensis” (Anaplasmataceae) is an emerging pathogen transmitted by Ixodes ticks. Conventional PCR and the newly developed high-resolution melt PCR were used to detect and discriminate “Candidatus Neoehrlichia mikurensis” and Anaplasma phagocytophilum. Both bacterial species were frequently found in Ixodes ricinus and Ixodes hexagonus but virtually absent from Dermacentor reticulatus. In rodents, “Candidatus N. mikurensis” was significantly more prevalent than A. phagocytophilum, whereas in cats, only A. phagocytophilum was found.
Candidatus Neoehrlichia mikurensis is a newly emerging tick-borne bacterium from the family Anaplasmataceae. Its presence in Ixodes ricinus ticks was reported from various European countries, however, it’s ecology and co-circulation with another member of the same family, Anaplasma phagocytophilum has not been rigorously studied yet.
Candidatus N. mikurensis was detected in all sampling sites. In total, 4.5% of ticks were positive including larvae. The highest positivity was detected in Austria with a prevalence of 23.5%. The probability of Candidatus N. mikurensis occurrence increased with the proportion of ticks infected with Anaplasma phagocytophilum.
A positive association between the occurrences of Candidatus N. mikurensis and A. phagocytophilum indicates that both bacteria share similar ecology for their natural foci in Central Europe.
Candidatus Neoehrlichia mikurensis; Anaplasma phagocytophilum; Ixodes ricinus; Human granulocytic anaplasmosis; Neoehrlichiosis
An immunocompromised patient presented with febrile episodes, an erysipelas-like rash, and thromboembolic complications. Amplification of 16S rRNA gene sequences from blood and sequence analysis revealed “Candidatus Neoehrlichia mikurensis.” We report the first case of human disease caused by “Ca. Neoehrlichia mikurensis.”
“Candidatus Neoehrlichia mikurensis” was detected by PCR in 4.0% (34/841) of the rodents tested in this study. The 34 rodents represented nine species from seven regions of China. Phylogenetic analyses based on the partial groEL and nearly entire 16S rRNA gene sequences of the agent revealed genetic diversity, which was correlated with its geographic origins.
We have repeatedly detected Candidatus Neoehrlichia mikurensis, a bacterium first described in Rattus norvegicus rats and Ixodes ovatus ticks in Japan in 2004 in the blood of a 61-year-old man with signs of septicemia by 16S rRNA and groEL gene PCR. After 6 weeks of therapy with doxycycline and rifampin, the patient recovered.
Candidatus Neoehrlichia mikurensis; septicemia; human infection; 16S rRNA gene PCR; therapy; tick-borne pathogen; bacteria; dispatch
“Candidatus Neoehrlichia mikurensis” is a new intracellular pathogen associated with human infection and death. “Candidatus Neoehrlichia mikurensis” infection in a chronically neutropenic dog from Germany was confirmed by DNA sequencing. The same organism was previously described from ticks and two sick human beings from Germany.
Altogether 2004 Ixodes ricinus ticks, from 37 places in Hungary, were analysed in pools with a recently developed multiplex real-time PCR for the presence of Candidatus Neoehrlichia mikurensis and for other representatives of the genus. Ca. Neoehrlichia mikurensis was identified in nine sampling sites, indicating three separated endemic regions along the borders of Hungary. In addition, results of samples from seven places (except for the western part of the country) were positive in the genus-specific (Ca. Neoehrlichia sp.) PCR, but were negative for Ca. Neoehrlichia mikurensis.
Tick-borne diseases; Zoonosis; Epidemiology
We examined small mammals as hosts for Anaplasmataceae in southern Sweden. Of 771 rodents, 68 (8.8%) were infected by Candidatus Neoehrlichia mikurensis, but no other Anaplasmataceae were found. Candidatus N. mikurensis has recently been found in human patients in Germany, Switzerland, and Sweden, which suggests that this could be an emerging pathogen in Europe.
Anaplasmataceae; Candidatus Neoehrlichia mikurensis; Anaplasma phagocytophilum; Bartonella; bacteria; tick-borne disease; zoonotic disease; wild rodents; Rickettsia; Sweden; dispatch
To estimate the likelihood of people coming into contact with the recently described tick-borne agent “Candidatus Neoehrlichia mikurensis,” we compared its prevalence to those of Lyme disease spirochetes and Anaplasma phagocytophilum in questing adult Ixodes ricinus ticks collected in various Central European sites and examined ticks, which had been removed from people, for the presence of these pathogens. Whereas spirochetes infected questing adult ticks most frequently (22.3%), fewer than a third as many ticks were infected by “Ca. Neoehrlichia mikurensis” (6.2%), and about a sixth harbored A. phagocytophilum (3.9%). On average, every twelfth encounter of a person with an I. ricinus tick (8.1%) may bear the risk of acquiring “Ca. Neoehrlichia mikurensis.” Although a fifth of the people (20%) had removed at least one tick infected by “Ca. Neoehrlichia mikurensis,” none displayed symptoms described for this pathogen, suggesting that its transmission may not be immediate and/or that immunocompetent individuals may not be affected. Because immunosuppressed patients may be at a particular risk of developing symptoms, it should be considered that “Ca. Neoehrlichia mikurensis” appears to be the second most common pathogen in I. ricinus ticks. In our survey, only Borrelia afzelii appears to infect Central European vector ticks more frequently.
To identify Candidatus Neoehrlichia mikurensis infection in northeastern China, we tested blood samples from 622 febrile patients. We identified in 7 infected patients and natural foci for this bacterium. Field surveys showed that 1.6% of ticks and 3.8% of rodents collected from residences of patients were also infected.
Candidatus Neoehrlichia mikurensis; bacteria; human infection; ticks; rodents; vector-borne infections; China
In Europe, Ixodes ricinus is the vector of many pathogens of medical and veterinary relevance, among them Borrelia burgdorferi sensu lato and tick-borne encephalitis virus, which have been the subject of numerous investigations. Less is known about the occurrence of emerging tick-borne pathogens like Rickettsia spp., Babesia spp., “Candidatus Neoehrlichia mikurensis,” and Anaplasma phagocytophilum in questing ticks. In this study, questing nymph and adult I. ricinus ticks were collected at 11 sites located in Western Switzerland. A total of 1,476 ticks were analyzed individually for the simultaneous presence of B. burgdorferi sensu lato, Rickettsia spp., Babesia spp., “Candidatus Neoehrlichia mikurensis,” and A. phagocytophilum. B. burgdorferi sensu lato, Rickettsia spp., and “Candidatus Neoehrlichia mikurensis” were detected in ticks at all sites with global prevalences of 22.5%, 10.2%, and 6.4%, respectively. Babesia- and A. phagocytophilum-infected ticks showed a more restricted geographic distribution, and their prevalences were lower (1.9% and 1.5%, respectively). Species rarely reported in Switzerland, like Borrelia spielmanii, Borrelia lusitaniae, and Rickettsia monacensis, were identified. Infections with more than one pathogenic species, involving mostly Borrelia spp. and Rickettsia helvetica, were detected in 19.6% of infected ticks. Globally, 34.2% of ticks were infected with at least one pathogen. The diversity of tick-borne pathogens detected in I. ricinus in this study and the frequency of coinfections underline the need to take them seriously into consideration when evaluating the risks of infection following a tick bite.
Candidatus Neoehrlichia mikurensis (CNM) has been described in the hard tick Ixodes ricinus and rodents as well as in some severe cases of human disease. The aims of this study were to identify DNA of CNM in small mammals, the ticks parasitizing them and questing ticks in areas with sympatric existence of Ixodes ricinus and Dermacentor reticulatus in Germany.
Blood, transudate and organ samples (spleen, kidney, liver, skin) of 91 small mammals and host-attached ticks from altogether 50 small mammals as well as questing I. ricinus ticks (n=782) were screened with a real-time PCR for DNA of CNM.
52.7% of the small mammals were positive for CNM-DNA. The majority of the infected animals were yellow-necked mice (Apodemus flavicollis) and bank voles (Myodes glareolus). Small mammals with tick infestation were more often infected with CNM than small mammals without ticks. Compared with the prevalence of ~25% in the questing I. ricinus ticks, twice the prevalence in the rodents provides evidence for their role as reservoir hosts for CNM.
The high prevalence of this pathogen in the investigated areas in both rodents and ticks points towards the need for more specific investigation on its role as a human pathogen.
Candidatus Neoehrlichia mikurensis; Bank vole; Yellow-necked mouse; Ixodes ricinus; Dermacentor reticulatus; Recreational area; Host survey; Vector-host relation
Ixodes ricinus, a competent vector of several pathogens, is the tick species most frequently reported to bite humans in Europe. The majority of human cases of Lyme borreliosis (LB) and tick-borne encephalitis (TBE) occur in the north-eastern region of Italy. The aims of this study were to detect the occurrence of endemic and emergent pathogens in north-eastern Italy using adult tick screening, and to identify areas at risk of pathogen transmission. Based on our results, different strategies for tick collection and pathogen screening and their relative costs were evaluated and discussed.
From 2006 to 2008 adult ticks were collected in 31 sites and molecularly screened for the detection of pathogens previously reported in the same area (i.e., LB agents, TBE virus, Anaplasma phagocytophilum, Rickettsia spp., Babesia spp., "Candidatus Neoehrlichia mikurensis"). Based on the results of this survey, three sampling strategies were evaluated a-posteriori, and the impact of each strategy on the final results and the overall cost reductions were analyzed. The strategies were as follows: tick collection throughout the year and testing of female ticks only (strategy A); collection from April to June and testing of all adult ticks (strategy B); collection from April to June and testing of female ticks only (strategy C).
Eleven pathogens were detected in 77 out of 193 ticks collected in 14 sites. The most common microorganisms detected were Borrelia burgdorferi sensu lato (17.6%), Rickettsia helvetica (13.1%), and "Ca. N. mikurensis" (10.5%). Within the B. burgdorferi complex, four genotypes (i.e., B. valaisiana, B. garinii, B. afzelii, and B. burgdorferi sensu stricto) were found. Less prevalent pathogens included R. monacensis (3.7%), TBE virus (2.1%), A. phagocytophilum (1.5%), Bartonella spp. (1%), and Babesia EU1 (0.5%). Co-infections by more than one pathogen were diagnosed in 22% of infected ticks. The prevalences of infection assessed using the three alternative strategies were in accordance with the initial results, with 13, 11, and 10 out of 14 sites showing occurrence of at least one pathogen, respectively. The strategies A, B, and C proposed herein would allow to reduce the original costs of sampling and laboratory analyses by one third, half, and two thirds, respectively. Strategy B was demonstrated to represent the most cost-effective choice, offering a substantial reduction of costs, as well as reliable results.
Monitoring of tick-borne diseases is expensive, particularly in areas where several zoonotic pathogens co-occur. Cost-effectiveness studies can support the choice of the best monitoring strategy, which should take into account the ecology of the area under investigation, as well as the available budget.
Ixodes ricinus; tick-borne diseases; surveillance; economic evaluation; Italy.
DNA microarrays can be used to quickly and sensitively identify several different pathogens in one step. Our previously developed DNA microarray, based on the detection of variable regions in the 16S rDNA gene (rrs), which are specific for each selected bacterial genus, allowed the concurrent detection of Borrelia spp., Anaplasma spp., Francisella spp., Rickettsia spp. and Coxiella spp.
In this study, we developed a comprehensive detection system consisting of a second generation DNA microarray and quantitative PCRs. New oligonucleotide capture probes specific for Borrelia burgdorferi s.l. genospecies and Candidatus Neoehrlichia mikurensis were included. This new DNA microarray system required substantial changes in solution composition, hybridization conditions and post-hybridization washes.
This second generation chip displayed high specificity and sensitivity. The specificity of the capture probes was tested by hybridizing the DNA microarrays with Cy5-labeled, PCR-generated amplicons encoding the rrs genes of both target and non-target bacteria. The detection limit was determined to be 103 genome copies, which corresponds to 1–2 pg of DNA. A given sample was evaluated as positive if its mean fluorescence was at least 10% of the mean fluorescence of a positive control. Those samples with fluorescence close to the threshold were further analyzed using quantitative PCRs, developed to identify Francisella spp., Rickettsia spp. and Coxiella spp. Like the DNA microarray, the qPCRs were based on the genus specific variable regions of the rrs gene. No unspecific cross-reactions were detected. The detection limit for Francisella spp. was determined to be only 1 genome copy, for Coxiella spp. 10 copies, and for Rickettsia spp., 100 copies.
Our detection system offers a rapid method for the comprehensive identification of tick-borne bacteria, which is applicable to clinical samples. It can also be used to identify both pathogenic and endosymbiontic bacteria in ticks for eco-epidemiological studies, tick laboratory colony testing, and many other applications.
Tick-borne bacteria; DNA microarray; Quantitative PCR
To further assess the geographic occurrence, possible vectors, and prevalence of Candidatus Neoehrlichia mikurensis, we analyzed spleen tissues from 276 voles trapped close to human settlements in France; 5 were infected with the organism. Sequencing showed the isolates carried the same genotype as the bacteria that caused disease in humans and animals elsewhere in Europe.
Candidatus Neoehrlichia mikurensis; rodents; bank vole; Myodes glareolus; France; zoonoses; wildlife; vector-borne infections; ticks; Ixodes ricinus; bacteria
Raising abundance of ticks and tick-borne diseases in Europe is the result of multiple factors including climate changes and human activities. Herein, we investigated the presence and seasonal activity of Ixodes ricinus ticks from 10 urban and suburban sites in two different geographical areas of southeastern and northeastern Slovakia during 2008–2010. Our aim was to study the abundance of ticks in correlation with the environmental factors and their infection with Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum and Neoehrlichia mikurensis.
Questing I. ricinus ticks were collected from ten urban and suburban sites in Eastern Slovakia. A total of 670 ticks were further analysed for the presence of B. burgdorferi s.l., A. phagocytophilum and N. mikurensis by molecular methods. Tick site and environmental relations were analysed using General Linear Models (LM). The differences between the number of Lyme borreliosis cases between the Košice and Bardejov regions during a ten-year period were tested by Wilcoxon matched pairs test.
In total, 2921 (1913 nymphs, 1008 adults) I. ricinus ticks were collected from 10 study sites during the main questing season. Tick activity and relative abundance differed between locations and months. Temperature and humidity were the main factors affecting the tick abundance and questing activity. Out of 670 examined ticks, 10.15% were infected with spirochetes from B. burgdorferi s.l. complex (represented by B. afzelii, B. garinii, B.valaisiana and B. burgdorferi s.s.), 2.69% with the A. phagocytophilum and 2.39% with N. mikurensis. The number of Lyme borreliosis cases per 100,000 inhabitants in the Bardejov region was significantly higher than in the Košice region.
Our data indicate that the risk of infection with tick-borne pathogens in Eastern Slovakia is common since 15.2% of ticks were infected at least with one of the tested microorganisms. Even though the abundance of ticks was affected by the microclimatic conditions and the prevalence of pathogens differed between the habitats, the infection risk for humans is also affected by human activities leading to an increased contact with infected ticks.
Ixodes ricinus; Borrelia burgdorferi sensu lato; Anaplasma phagocytophilum; Neoehrlichia mikurensis; PCR-RFLP; Lyme borreliosis; Anaplasmosis
Members of the uncultured bacterial genus Candidatus Accumulibacter are capable of intracellular accumulation of inorganic phosphate (Pi) in activated sludge wastewater treatment plants (WWTPs) performing enhanced biological phosphorus removal (EBPR), but were also recently shown to inhabit freshwater and estuarine sediments. Additionally, metagenomic sequencing of two bioreactor cultures enriched in Candidatus Accumulibacter, but housed on separate continents, revealed the potential for global dispersal of particular Candidatus Accumulibacter strains, that we hypothesize is facilitated by the ability of Candidatus Accumulibacter to persist in environmental habitats. In the current study, we used sequencing of a phylogenetic marker, the ppk1 gene, to characterize Candidatus Accumulibacter populations in diverse environments, at varying distances from WWTPs. We discovered several new lineages of Candidatus Accumulibacter which had not previously been detected in WWTPs, and also uncovered new diversity and structure within previously detected lineages. Habitat characteristics were found to be a key determinant of Candidatus Accumulibacter lineage distribution, while, as predicted, geographic distance played little role in limiting dispersal on a regional scale. However, on a local scale, enrichment of particular Candidatus Accumulibacter lineages in WWTP appeared to impact local environmental populations. These results provide evidence of ecological differences among Candidatus Accumulibacter lineages.
The order Rickettsiales (Alphaproteobacteria) is a well-known group containing obligate endocellular prokaryotes. The order encompasses three families (Rickettsiaceae, Anaplasmataceae, and Holosporaceae) and a fourth, family-level cluster, which includes only one candidate species, “Candidatus Midichloria mitochondrii,” as well as several unnamed bacterial symbionts. The broad host range exhibited by the members of the “Candidatus Midichloria” clade suggests their eventual relevance for a better understanding of the evolution of symbiosis and host specificity of Rickettsiales. In this paper, two new bacteria belonging to the “Candidatus Midichloria” clade, hosted by two different strains of the ciliate protist Euplotes harpa, are described on the basis of ultrastructural observations, comparative 16S rRNA gene sequence analysis, and an estimation of the percentage of infection. Ultrastructure of these bacteria shows some unusual features: one has an electron-dense cytoplasm, and the other one lacks a symbiosomal membrane. The latter was up to now considered an exclusive feature of bacteria belonging to the family Rickettsiaceae. 16S rRNA gene phylogenetic analysis unambiguously places the new bacteria in the “Candidatus Midichloria” clade, although their phylogenetic relationships with other members of the clade are not clearly resolved. This is the first report of a ciliate-borne bacterium belonging to the “Candidatus Midichloria” clade. On the basis of the data obtained, the two bacteria are proposed as two new candidate genera and species, “Candidatus Anadelfobacter veles” and “Candidatus Cyrtobacter comes.”
Ixodes ricinus transmits Borrelia burgdorferi sensu lato, the etiological agent of Lyme disease. Previous studies have also detected Rickettsia helvetica, Anaplasma phagocytophilum, Neoehrlichia mikurensis, and several Babesia species in questing ticks in The Netherlands. In this study, we assessed the acarological risk of exposure to several tick-borne pathogens (TBPs), in The Netherlands. Questing ticks were collected monthly between 2006 and 2010 at 21 sites and between 2000 and 2009 at one other site. Nymphs and adults were analysed individually for the presence of TBPs using an array-approach. Collated data of this and previous studies were used to generate, for each pathogen, a presence/absence map and to further analyse their spatiotemporal variation. R. helvetica (31.1%) and B. burgdorferi sensu lato (11.8%) had the highest overall prevalence and were detected in all areas. N. mikurensis (5.6%), A. phagocytophilum (0.8%), and Babesia spp. (1.7%) were detected in most, but not all areas. The prevalences of pathogens varied among the study areas from 0 to 64%, while the density of questing ticks varied from 1 to 179/100 m2. Overall, 37% of the ticks were infected with at least one pathogen and 6.3% with more than one pathogen. One-third of the Borrelia-positive ticks were infected with at least one other pathogen. Coinfection of B. afzelii with N. mikurensis and with Babesia spp. occurred significantly more often than single infections, indicating the existence of mutual reservoir hosts. Alternatively, coinfection of R. helvetica with either B. afzelii or N. mikurensis occurred significantly less frequent. The diversity of TBPs detected in I. ricinus in this study and the frequency of their coinfections with B. burgdorferi s.l., underline the need to consider them when evaluating the risks of infection and subsequently the risk of disease following a tick bite.
vector-borne disease; Borrelia burgdorferi; Candidatus Neoehrlichia mikurensis; Rickettsia helvetica; Rickettsia conorii; Anaplasma phagocytophilum; Babesia; Ixodes ricinus
Kinetic analyses of GroE-assisted folding provide a dynamic sequence of molecular events that underlie chaperonin function. We used stopped-flow analysis of various fluorescent GroEL mutants to obtain details regarding the sequence of events that transpire immediately after ATP binding to GroEL and GroEL with prebound unfolded proteins. Characterization of GroEL CP86, a circularly permuted GroEL with the polypeptide ends relocated to the vicinity of the ATP binding site, showed that GroES binding and protection of unfolded protein from solution is achieved surprisingly early in the functional cycle, and in spite of greatly reduced apical domain movement. Analysis of fluorescent GroEL SR-1 and GroEL D398A variants suggested that among other factors, the presence of two GroEL rings and a specific conformational rearrangement of Helix M in GroEL contribute significantly to the rapid release of unfolded protein from the GroEL apical domain.
Three hemotropic mycoplasmas have been identified in pet cats: Mycoplasma haemofelis, “Candidatus Mycoplasma haemominutum,” and “Candidatus Mycoplasma turicensis.” The way in which these agents are transmitted is largely unknown. Thus, this study aimed to investigate fleas, ticks, and rodents as well as saliva and feces from infected cats for the presence of hemotropic mycoplasmas, to gain insight into potential transmission routes for these agents. DNA was extracted from arthropods and from rodent blood or tissue samples from Switzerland and from salivary and fecal swabs from two experimentally infected and six naturally infected cats. All samples were analyzed with real-time PCR, and some positive samples were confirmed by sequencing. Feline hemotropic mycoplasmas were detected in cat fleas and in a few Ixodes sp. and Rhipicephalus sp. ticks collected from animals but not in ticks collected from vegetation or from rodent samples, although the latter were frequently Mycoplasma coccoides PCR positive. When shedding patterns of feline hemotropic mycoplasmas were investigated, “Ca. Mycoplasma turicensis” DNA was detected in saliva and feces at the early but not at the late phase of infection. M. haemofelis and “Ca. Mycoplasma haemominutum” DNA was not amplified from saliva and feces of naturally infected cats, despite high hemotropic mycoplasma blood loads. Our results suggest that besides an ostensibly indirect transmission by fleas, direct transmission through saliva and feces at the early phase of infection could play a role in the epizootiology of feline hemotropic mycoplasmas. Neither the investigated tick nor the rodent population seems to represent a major reservoir for feline hemotropic mycoplasmas in Switzerland.
Recently, a third novel feline hemotropic Mycoplasma sp. (aka hemoplasma), “Candidatus Mycoplasma turicensis,” in a cat with hemolytic anemia has been described. This is the first study to investigate the prevalence, clinical manifestations, and risk factors for all three feline hemoplasma infections in a sample of 713 healthy and ill Swiss cats using newly designed quantitative real-time PCR assays. “Candidatus Mycoplasma haemominutum” infection was detected in 7.0% and 8.7% and Mycoplasma haemofelis was detected in 2.3% and 0.2% of healthy and ill cats, respectively. “Candidatus Mycoplasma turicensis” was only detected in six ill cats (1.1%); three of them were coinfected with “Candidatus Mycoplasma haemominutum.” The 16S rRNA gene sequence of 12 Swiss hemoplasma isolates revealed >98% similarity with previously published sequences. Hemoplasma infection was associated with male gender, outdoor access, and old age but not with retrovirus infection and was more frequent in certain areas of Switzerland. “Candidatus Mycoplasma haemominutum”-infected ill cats were more frequently diagnosed with renal insufficiency and exhibited higher renal blood parameters than uninfected ill cats. No correlation between hemoplasma load and packed cell volume was found, although several hemoplasma-infected cats, some coinfected with feline immunodeficiency virus or feline leukemia virus, showed hemolytic anemia. High M. haemofelis loads (>9 × 105 copies/ml blood) seem to lead to anemia in acutely infected cats but not in recovered long-term carriers. A repeated evaluation of 17 cats documented that the infection was acquired in one case by blood transfusion and that there were important differences among species regarding whether or not antibiotic administration led to the resolution of bacteremia.