Phylogenetic relationships among species of the genus Oligoryzomys (Rodentia,Cricetidae) from Central and South America

González‐Ittig, R. E., Salazar‐Bravo, J., Barquez, R. M. & Gardenal, C. N. (2010). Phylogenetic relationships among species of the genus Oligoryzomys (Rodentia, Cricetidae) from Central and South America. —Zoologica Scripta, 39, 511–526. The genus Oligoryzomys includes several species very similar in external morphology, which has resulted in a confusing specific taxonomy. Accurate species identification is particularly important because several species of Oligoryzomys act as natural hosts of hantaviruses affecting humans. Here, we assign specific status to individuals from a wide geographical area of Argentina and Chile using sequences of the mtDNA control region. We also compare cytochrome b sequences of 14 species recognized from Central and South America to infer the phylogenetic relationships within the genus. In addition, the results were analysed using available data on chromosome numbers, and the host–parasite relationships reported for the genus Hantavirus. We confirm the geographical distribution of Oligoryzomys longicaudatus (Argentina, Chile), Oligoryzomys nigripes (Argentina, Paraguay, Brazil), Oligoryzomys chacoensis (Argentina, Bolivia, Paraguay), Oligoryzomys fornesi (Argentina, Paraguay), Oligoryzomys destructor (Argentina, Bolivia) and Oligoryzomys microtis (Bolivia, Brazil). Oligoryzomys longicaudatus is strongly related to the Oligoryzomys flavescens complex, which comprises four clades; O. nigripes is closely related to Oligoryzomys stramineus, and Oligoryzomys vegetus, to Oligoryzomys fulvescens from Central America. Oligoryzomys chacoensis, O. destructor, O. fornesi, O. longicaudatus, O. microtis, O. nigripes, O. stramineus, Oligoryzomys moojeni, Oligoryzomys rupestris, O. fulvescens and O. vegetus are confirmed as valid species, whereas O. flavescens, Oligoryzomys magellanicus, Oligoryzomys griseolus, Oligoryzomys victus, Oligoryzomys andinus and Oligoryzomys arenalis need exhaustive revision. The sister species to all the remaining entities of the genus was O. microtis, suggesting an Amazonian origin for the genus.     

Diversity and Distribution of Hantaviruses in South America

Hantaviruses are important contributors to disease burden in the New World, yet many aspects of their distribution and dynamics remain uncharacterized. To examine the patterns and processes that influence the diversity and geographic distribution of hantaviruses in South America, we performed genetic and phylogeographic analyses of all available South American hantavirus sequences. We sequenced multiple novel and previously described viruses (Anajatuba, Laguna Negra-like, two genotypes of Castelo dos Sonhos, and two genotypes of Rio Mamore) from Brazilian Oligoryzomys rodents and hantavirus pulmonary syndrome cases and identified a previously uncharacterized species of Oligoryzomys associated with a new genotype of Rio Mamore virus. Our analysis indicates that the majority of South American hantaviruses fall into three phylogenetic clades, corresponding to Andes and Andes-like viruses, Laguna Negra and Laguna Negra-like viruses, and Rio Mamore and Rio Mamore-like viruses. In addition, the dynamics and distribution of these viruses appear to be shaped by both the geographic proximity and phylogenetic relatedness of their rodent hosts. The current system of nomenclature used in the hantavirus community is a significant impediment to understanding the ecology and evolutionary history of hantaviruses; here, we suggest strict adherence to a modified taxonomic system, with species and strain designations resembling the numerical system of the enterovirus genus.     

Temporal and Spatial Host Abundance and Prevalence of Andes Hantavirus in Southern Argentina

Andes virus (AND) is a hantavirus hosted by the sigmodontine rodent Oligoryzomys longicaudatus in southern Argentina, where it is responsible for most cases of hantavirus pulmonary syndrome (HPS). Our study provides data about the spatial variation in abundance of the rodent host of AND hantavirus. We report results of a longitudinal study performed in a locality of the Andean region of Chubut Province. From November 2003 (spring) to July 2006 (winter), O. longicaudatus was the most common species captured (63%) and it showed significant differences in abundance among habitats and seasons. Most antibody-positive rodents were O. longicaudatus (9.2%), followed by A. longipilis (3.6%) and A. olivaceus (1.5%). The highest number of antibody-positive animals was observed for males that belonged to the heaviest mass classes. Antibody-positive O. longicaudatus were more abundant in brush habitats. We found low richness of rodents and abundance of O. longicaudatus in areas affected by anthropogenic activity. The infection seems to be regionally persistent, but the risk to humans in a landscape would be localized. To develop accurate models for predicting HPS outbreaks, further research is needed to characterize rodent movement patterns across the landscape.     

Phylogenetic relationships of the pygmy rice rats of the genus Oligoryzomys Bangs, 1900 (Rodentia: Sigmodontinae)

Sequences from two mitochondrial genes (cytochrome b and NADH1) were used to produce a molecular phylogeny for 12 named and two undescribed species of the genus Oligoryzomys. All analyses placed Oligoryzomys microtis as the most basal taxon, a finding consistent with previous studies that suggested the west‐central Amazon as a centre of origin for the tribe Oryzomyini to which Oligoryzomys belongs. Biogeographically, this suggests that Oligoryzomys had a South American origin, and later advanced northwards, entering Central America and Mexico more recently. Different analyses have provided consistent support for several additional clades that did not necessarily agree with the species groups hypothesized by previous studies. A molecular clock derived for these data suggests an origin for the genus of 6.67 Mya, with most speciation within the genus occurring between 3.7 and 1.5 Mya. © 2010 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010, 160 , 551–566.     

Molecular identification and phylogenetic relationships among the species of the genus Oligoryzomys (Rodentia, Cricetidae) present in Argentina, putative reservoirs of hantaviruses

The systematics and geographical distribution of the species of Oligoryzomys present in Argentina are poorly known. From some of the species different hantavirus genotypes have been recovered. In order to contribute to the accurate identification of those species and to infer their phylogenetic relationships, we analysed data of restriction sites and sequences of the mtDNA d -loop region. The samples used represent almost all Oligoryzomys species known to occur in Argentina. The trees obtained with the two types of data were similar, showing high bootstrap values for the majority of the nodes. Our results support the idea that the specific name Oligoryzomys longicaudatus should be applied only to individuals from the south of Argentina and Chile and confirm that O. nigripes and O. delticola are conspecific. Specimens identified as O. flavescens conform three related clades, probably belonging to a species complex. This study also emphasises that the use of a DNA fragment characterised by a high evolutionary rate compare with other mitochondrial segments as the d -loop, was appropriate to infer the phylogenetic relationships in a group originated by a rapid speciation process. We also suggest the following modifications in the rodent–hantavirus relationships: the only viral genotype associated to O. longicaudatus would be Andes Sout, while O. chacoensis would be the natural host of the genotype Oran. One of the forms of the O. flavescens complex would be the natural host of the genotype Bermejo. Our data are consistent with the hypothesis that each hantavirus genotype is associated with a specific rodent host.     

Potential Geographic Distribution of Hantavirus Reservoirs in Brazil

Hantavirus cardiopulmonary syndrome is an emerging zoonosis in Brazil. Human infections occur via inhalation of aerosolized viral particles from excreta of infected wild rodents. Necromys lasiurus and Oligoryzomys nigripes appear to be the main reservoirs of hantavirus in the Atlantic Forest and Cerrado biomes. We estimated and compared ecological niches of the two rodent species, and analyzed environmental factors influencing their occurrence, to understand the geography of hantavirus transmission. N. lasiurus showed a wide potential distribution in Brazil, in the Cerrado, Caatinga, and Atlantic Forest biomes. Highest climate suitability for O. nigripes was observed along the Brazilian Atlantic coast. Maximum temperature in the warmest months and annual precipitation were the variables that most influence the distributions of N. lasiurus and O. nigripes, respectively. Models based on occurrences of infected rodents estimated a broader area of risk for hantavirus transmission in southeastern and southern Brazil, coinciding with the distribution of human cases of hantavirus cardiopulmonary syndrome. We found no demonstrable environmental differences among occurrence sites for the rodents and for human cases of hantavirus. However, areas of northern and northeastern Brazil are also apparently suitable for the two species, without broad coincidence with human cases. Modeling of niches and distributions of rodent reservoirs indicates potential for transmission of hantavirus across virtually all of Brazil outside the Amazon Basin.     

Comparative Chromosome Painting in Six Species of Oligoryzomys (Rodentia,Sigmodontinae) and the Karyotype Evolution of the Genus

Oligoryzomys belongs to the tribe Oryzomyini, and contains about 22 species. Diploid numbers range from 2n = 44 in Oligoryzomys sp. 2 to 2n = 72 in O. utiaritensis and phylogenetic relationships are not well defined. The high morphological convergence leads to misidentification of taxonomic entities and the species are often identified by chromosomal characters. Until now, the genus has been studied only by classical cytogenetic approaches. To understand the chromosomal evolution of Oligoryzomys, we developed chromosome probes from a female of Oligoryzomys moojeni (OMO) with 2n = 70 and hybridized to other five Oligoryzomys species. The probes painted 31 segments on O. fornesi (OFO) with 2n = 62; 32 segments on O. microtis (OMI), 2n = 64; 33 segments on O. nigripes (ONI), 2n = 62 and on O. rupestris (ORU), 2n = 46; and 34 on Oligoryzomys sp. 2 (OSP), 2n = 44. OMO probes 4 and 5 showed a syntenic association in O. fornesi, O. microtis and O. nigripes and were also presented in the same pair, although disrupted, in O. rupestris and Oligoryzomys sp. 2. Concerning O. rupestris and Oligoryzomys sp. 2, species with the lowest diploid numbers of the genus, a total of 8 probes hybridized to 11 segments on the largest pair of ORU 1 and 9 probes hybridized to 12 segments on OSP 1. Also, OMO 6 painted three segments in ORU, corresponding to the proximal segment of ORU 2q, and the whole of ORU 19 and 20. In OSP, the segment corresponding to ORU 20 was homologous to OSP 1p. OMO X showed signals of hybridization in both X and Y chromosomes. Extensive chromosomal rearrangements, that could not be detected by classical cytogenetic techniques, such as pericentric inversions or repositioning of centromeres, Robertsonian rearrangements and tandem fusions/fissions, as well as gain/activation or loss/inactivation of centromeres and telomeric sequences have driven the huge genome reshuffling in these closely related species.     

Taxonomy of the long‐tailed mouse Oligoryzomys destructor (Sigmodontinae: Oryzomyini) with the designation of neotypes for Hesperomys destructor Tschudi, 1844 and Hesperomys melanostoma Tschudi, 1844

The genus Oligoryzomys, distributed from southern South America to southern North America, is the most diverse of the tribe Oryzomyini of sigmodontine rodents. Even when 22 species are currently recognized, species boundaries are unclear for several forms. The species Oligoryzomys destructor is one of the least studied species of the genus and is the one with the largest distribution along the Andes (from southern Colombia to northern Bolivia). The species was described without the selection of a holotype and indication of its type locality. In addition, several taxa are regarded as synonyms of O. destructor. These facts are relevant because previous analysis of DNA sequences has shown that O. destructor represents a species complex. Herein, in addition to test the phylogenetic position of O. destructor within the genus Oligoryzomys, we assess patterns of morphological and molecular variation of O. destructor and its associated nominal forms aimed to assess the boundaries of the species. As part of the study, we selected neotypes for Hesperomys destructor and H. melanostoma. At the light of our results, we recognized O. destructor as a species with two subspecies, O. d. destructor and O. d. spodiurus. Also, we discuss the role of Andean rivers, and their different permeability, as allopatric barriers molding the structure of O. destructor.     

Long-Term Study of a Hantavirus Reservoir Population in an Urban Protected Area,Argentina

Green spaces in urban areas can play a key role in protecting wildlife. However, the presence of wildlife in urban areas can lead to human health risks. Although the presence of the rodent species Oligoryzomys flavescens (hantavirus reservoir) has been recorded in cities of Argentina, its population dynamics in this type of habitat is still unknown. Here, we evaluated: (1) long-term spatial and temporal patterns of O. flavescens abundance and how these patterns were influenced by weather factors and (2) the seroprevalence of hantavirus and the identity of the viral lineage circulating in the population that inhabits the Costanera Sur Ecological Reserve, a protected area in the city of Buenos Aires. Genetic results confirmed that the pathogenic ANDES Central Lechiguanas virus is present in O. flavescens populations inhabiting this urban reserve. Abundance of O. flavescens showed interannual and seasonal fluctuations, with maximum values in winter and spring and minimum ones in summer and autumn. Summers with the highest abundances of O. flavescens were preceded by warmer winters, while winters with lower abundances were preceded by warmer summers. On the other hand, accumulated precipitations in the previous 6 months positively affected winter abundance. These results could help the authorities in charge of the green spaces of Buenos Aires to identify priority areas and times of the year for the implementation of preventive measures that minimize the contact of rodents with visitors. Such measures could be intensified when winters are warmer than normal, and summers are cooler and wetter than normal.     

Space Use and Social Mating System of the Hantavirus Host, <Emphasis Type="Italic">Oligoryzomys longicaudatus</Emphasis>

The long-tailed mouse, Oligoryzomys longicaudatus (Cricetidae: Sigmodontinae), is the major host of Andes hantavirus, the etiological agent of hantavirus pulmonary syndrome in the south of Argentina and Chile. Studying the ecology of this species is necessary to understand how Andes hantavirus is maintained in nature. In this study, we examine the home range size and intra- and intersexual overlap degree of male and female O. longicaudatus in order to elucidate the mating system of this species. To our knowledge, this research provides the first documentation, obtained from a specific design, of spacing and mating systems in this species in Argentina. The study was conducted seasonally from April (autumn) 2012 to October (spring) 2013 in a shrubland habitat of Cholila, Andean region, Argentina. We studied spacing patterns using 59 and 51 home ranges established by adult males and females, respectively, in two 3.24 ha capture-marked and recapture grids. Significant differences between sexes in home range size and overlap degree were found. Male home ranges were always larger than those of females. We observed exclusive space use both among males and females (13.15 ± 18.67, and 3.60 ± 3.43%, respectively). Considering only those males that get access to receptive females (40%), average intersexual overlap value was about 30.82 ± 19.73%. Sexual differences in home range sizes and the spatial avoidance between breeding males, that would reflect intrasexual competition for receptive females, allows us to propose a polygynous mating system for O. longicaudatus.     

A long term study of small mammal populations in a Brazilian agricultural landscape

Long-term monitoring of small mammal populations is very important to understand the variations in temporal abundance on a large time scale, which are related to ecological, economic and epidemiological phenomena. The aim of this study is to monitor the populations of the marsupials Didelphis aurita and Philander frenatus and the rodents Nectomys squamipes, Akodon cursor and Oligorysomys nigripes in a locality of typical Brazilian rural landscape, Sumidouro Municipality, Rio de Janeiro State, Brazil. A mark-recapture study was conducted during five years. We analyzed the population dynamics, the reproduction and age structure of these species. Both marsupials presented higher population sizes in the end of wet period and beginning of the dry period, which can be explained by the seasonal reproduction which begins in the middle of the dry period and ends in the last months of the wet period. N. squamipes reproduced throughout the year but mostly during rainy periods, due to the close association of this rodent to resources found in the water. Higher survivorship and recruitment rates were in the end of the wet season. The rodent A. cursor had an opportunist reproduction, resulting in high turnover rates. Survivorship increased with the effects of the dry periods. O. nigripes showed a clear annual pattern of population cycle with peaks during the dry season. The rodents did not show potential to present outbreaks and become agricultural pests. The annual population cycles of O. nigripes and the unique peak of A. cursor population during five years highlight attention to their importance as wild reservoirs of the hantavirus disease. Their ecological characteristics associated to their opportunistic behavior make these species prone to be good reservoirs of zoonoses.     

Silent <Emphasis Type="Italic">Orthohantavirus</Emphasis> Circulation Among Humans and Small Mammals from Central Minas Gerais,Brazil

New World orthohantaviruses are emerging RNA viruses that cause hantavirus cardiopulmonary syndrome (HCPS). These viruses are a burden to public health around the world with a lethality rate of around 60%. In South America, rodents of Sigmodontinae subfamily are the main reservoirs of orthohantaviruses. We described a serosurvey for orthohantaviruses circulation in an apparently healthy human population and small mammals from rural areas in Central Minas Gerais State, Brazil. A total of 240 individuals and 50 small mammals (26 rodents belonging to 10 different species and 24 marsupials from 4 different species) were sampled during 2012–2013. The seroprevalence rates of IgG/IgM antibodies in humans were 7.1 and 1.6%, respectively. Only one rodent, an Oligoryzomys nigripes captured in peridomestic area, tested positive for IgG antibodies and viral RNA. Our findings suggest a silent circulation of orthohantaviruses in a region of intensive agriculture production. The detection of seropositive humans in an area with a lack of previous HCPS reports highlights potential oligosymptomatic cases and the need for surveillance strategies that could reduce the risk of future outbreaks.     

Experimental Andes Virus Infection in Deer Mice: Characteristics of Infection and Clearance in a Heterologous Rodent Host

New World hantaviruses can cause hantavirus cardiopulmonary syndrome with high mortality in humans. Distinct virus species are hosted by specific rodent reservoirs, which also serve as the vectors. Although regional spillover has been documented, it is unknown whether rodent reservoirs are competent for infection by hantaviruses that are geographically separated, and known to have related, but distinct rodent reservoir hosts. We show that Andes virus (ANDV) of South America, carried by the long tailed pygmy rice rat (Oligoryzomys longicaudatus), infects and replicates in vitro and in vivo in the deer mouse (Peromyscus maniculatus), the reservoir host of Sin Nombre virus (SNV), found in North America. In experimentally infected deer mice, viral RNA was detected in the blood, lung, heart and spleen, but virus was cleared by 56 days post inoculation (dpi). All of the inoculated deer mice mounted a humoral immune response by 14 dpi, and produced measurable amounts of neutralizing antibodies by 21 dpi. An up-regulation of Ccl3, Ccl4, Ccl5, and Tgfb, a strong CD4⁺ T-cell response, and down-regulation of Il17, Il21 and Il23 occurred during infection. Infection was transient with an absence of clinical signs or histopathological changes. This is the first evidence that ANDV asymptomatically infects, and is immunogenic in deer mice, a non-natural host species of ANDV. Comparing the immune response in this model to that of the immune response in the natural hosts upon infection with their co-adapted hantaviruses may help clarify the mechanisms governing persistent infection in the natural hosts of hantaviruses.     

A contribution to the subgeneric systematics of Oligoryzomys (Rodentia,Muridae) from Argentina by means of PCR–RFLP patterns of mitochondrial DNA

Subgeneric recognition of rodents of the genus Oligoryzomys from Argentina is specially difficult because morphologic and morphometric criteria to identify species are poorly defined. In order to contribute to the unequivocal identification and geographic distribution of Oligoryzomys species, we studied the restriction fragment length polymorphism (PCR-RFLP) patterns of the D-loop region of the mitochondrial DNA in specimens collected from different regions of Argentina. PCR amplification products of 59 individuals were digested with five restriction enzymes. Patterns obtained were used to produce two phylogenetic consensus trees, one obtained with PAUP and the other with MIX analyses. All specimens from the Patagonia (southern region) clustered together and would correspond to O. longicaudatus. Individuals from the North grouped in four clusters (two of them comprising one individual), supported by high bootstrap values. The one including the 82% of the rodents from the northern region would correspond to the species O. chacoensis.     

Ogmogaster heptalineatus n.sp. (Trematoda: Notocotylidae) from the Chilean sea lion Otaria flavescens

Summary Ogmogaster heptalineatus n.sp. is described from the Chilean sea lion Otaria flavescens. Distinct characteristics of the species are: presence of 7 longitudinal ridges on the ventral surface, cirrus armed with scales, testes and ovary deeply lobed and eggs circular with short polar filaments. This is the first report of the genus Ogmogaster from South America and the first report of Otaria flavescens as a host for this genus. ac]19820320     

Determinants of ectoparasite assemblage structure on rodent hosts from South American marshlands: the effect of host species,locality and season

The relative effects of host species identity, locality and season on ectoparasite assemblages (relative abundances and species richness) harboured by four cricetid rodent hosts (Akodon azarae, Oligoryzomys flavescens, Oxymycterus rufus and Scapteromys aquaticus) were assessed across six closely located sites in Buenos Aires province, Argentina. Relative abundances of ectoparasites (14 species including gamasid mites, an ixodid tick, a trombiculid mite, lice and fleas), as well as total ectoparasite abundance and species richness, were determined mainly by host species and to a lesser extent by locality (despite the small spatial scale of the study), whereas seasonal effect was weak, albeit significant. The abundances of some ectoparasites were determined solely by host, whereas those of other ectoparasites (sometimes belonging to the same higher taxon) were also affected by locality and/or season. In gamasids, there was a significant effect of locality for some species, but not for others. In fleas and lice, the effect of locality was similar in different species, suggesting that this effect is related to the characteristic life history strategy.     

An assessment of species limits of the South American mouse genus Oligoryzomys (Rodentia,Cricetidae) using unilocus delimitation methods

Oligoryzomys, as currently understood is formed by 25 living species, is the most diverse genus of the tribe Oryzomyini of the New World subfamily Sigmodontinae of cricetid rodents. Nonetheless, the species richness of Oligoryzomys seems to be an underestimate, given some species complex has been proposed in previous studies, at the time that large geographic areas remain to be sampled, and several taxonomic forms have not been assessed with contemporary approaches. In this study, we present a new assessment of the species diversity of Oligoryzomys based on multiple unilocus species delimitation methods (ABGD, BPP, PTP, GMYC and b GMYC), using 665 cytb gene sequences as evidence (532 gathered from Genbank and 133 obtained in this study). We sampled representatives of almost all currently known species of Oligoryzomys, at the time that extending the geographic coverage to the Central Andes, a large area that was largely unrepresented in previous studies. Phylogenetic relationships, based on a non‐redundant alignment, were inferred via maximum likelihood and Bayesian inference; an ultrametric tree, used in species delimitation analyses, was obtained using multiple secondary calibration points. Results of species delimitation methods are discussed at the light of previous knowledge (e.g., taxonomic history and geographic provenance of samples in relation to type localities) and the morphological assessments of some specimens. Results of the distinct delimitation methods are mostly congruent, being BPP and PTP the most sensible to estimate species delimitation, allowing us to suggest that Oligoryzomys is composed of 30 lineages of species level. Of these, 22 correspond to forms currently considered species; some of these include in their synonymy some forms currently considered valid species (e.g., yatesi would be a synonym of longicaudatus). The remaining eight lineages are candidate species that need to be further evaluated. This study, by advancing taxonomic hypothesis that should be further tested in future studies, constitutes a stepping‐stone for upcoming taxonomic and biogeographic studies centred on Oligoryzomys.     

Isolation and characterization of microsatellite markers in Oligoryzomys longicaudatus (Muridae, Sigmodontinae, Oryzomini), the natural reservoir of genotype Andes hantavirus

The rodent Oligoryzomys longicaudatus or long-tailed pygmy rice rat is the reservoir of the aetiological agent of the hantavirus pulmonary syndrome in southern Argentina and Chile. We characterize 11 polymorphic microsatellite loci which would be useful for studies on microgeographical population structure in the species. Amplification of these loci in 42 individuals from four natural populations revealed four to 21 alleles per locus, and values of observed heterozygosities ranging from 0.371 to 0.896. Cross-species amplifications showed that some of the primers designed may be useful for other species of the genus Oligoryzomys.     

Molecular phylogeny and host-plant use (Lamiaceae) of the Thymogethes pollen beetles (Coleoptera)

The 24 members of the Euro-Asiatic genus Thymogethes are highly specialized pollen beetles associated as larvae with flowers of Lamiaceae Nepetoideae. All members of the genus were analysed in within the framework of an integrative taxonomy approach, which was aimed to reconstruct the phylogenetic relationships, as well as the possible pattern of evolution of their larval-host-plant association. Evidence from multiple molecular markers [COI; 16S; H3], combined with an estimation of divergence times using an average rate of 0.0177 substitutions/site/My among branches, placed the origin of the genus at a minimum of 9–10 Mya. This date of origin approximates the known evolution of the host plants in Euro-Mediterranean areas. Evidence from combined molecular and cladistic morphological analyses resulted in suitable agreement with the previously established morphology-based systematics of the genus, although members of the exilis species-group were split into three clades. The only disagreement between results of this new combined phylogeny and previous classification is in the exclusion of “Thymogethes” grenieri. This species is herein positioned outside the genus, based on molecular evidence. Our analysis depicts several Thymogethes species differentiating in the last few Mys, specifically those included in the T. lugubris species-group. Combined evidence from DNA, morphology and ancestral state parsimony reconstruction of larval-host-plant associations suggests that subtribe Menthinae likely represents the ancestral host plants, with a series of independent host shifts during the radiation of the clade, in association first with Menthinae and subsequently with Lavandulinae and Nepetinae. Steno-oligophagy is the most frequent (86%) condition, while strictly monophagous species are less numerous (14%).