Cancer-relevant Splicing Factor CAPERα Engages the Essential Splicing Factor SF3b155 in a Specific Ternary Complex

U2AF homology motifs (UHMs) mediate protein-protein interactions with U2AF ligand motifs (ULMs) of pre-mRNA splicing factors. The UHM-containing alternative splicing factor CAPERα regulates splicing of tumor-promoting VEGF isoforms, yet the molecular target of the CAPERα UHM is unknown. Here we present structures of the CAPERα UHM bound to a representative SF3b155 ULM at 1.7 Å resolution and, for comparison, in the absence of ligand at 2.2 Å resolution. The prototypical UHM/ULM interactions authenticate CAPERα as a bona fide member of the UHM family of proteins. We identify SF3b155 as the relevant ULM-containing partner of full-length CAPERα in human cell extracts. Isothermal titration calorimetry comparisons of the purified CAPERα UHM binding known ULM-containing proteins demonstrate that high affinity interactions depend on the presence of an intact, intrinsically unstructured SF3b155 domain containing seven ULM-like motifs. The interplay among bound CAPERα molecules gives rise to the appearance of two high affinity sites in the SF3b155 ULM-containing domain. In conjunction with the previously identified, UHM/ULM-mediated complexes of U2AF⁶⁵ and SPF45 with SF3b155, this work demonstrates the capacity of SF3b155 to offer a platform for coordinated recruitment of UHM-containing splicing factors.     

MiR-17 Partly Promotes Hematopoietic Cell Expansion through Augmenting HIF-1α in Osteoblasts

Background

Hematopoietic stem cell (HSC) regulation is highly dependent on interactions with the marrow microenvironment, of which osteogenic cells play a crucial role. While evidence is accumulating for an important role of intrinsic miR-17 in regulating HSCs and HPCs, whether miR-17 signaling pathways are also necessary in the cell-extrinsic control of hematopoiesis hereto remains poorly understood.

Methodology/Principal Findings

Using the immortalized clone with the characteristics of osteoblasts, FBMOB-hTERT, in vitro expansion, long-term culture initiating cell (LTC-IC) and non-obese diabetic/severe combined immunodeficient disease (NOD/SCID) mice repopulating cell (SRC) assay revealed that the ectopic expression of miR-17 partly promoted the ability of FBMOB-hTERT to support human cord blood (CB) CD34⁺ cell expansion and maintain their multipotency. It also seemed that osteoblastic miR-17 was prone to cause a specific expansion of the erythroid lineage. Conversely, deficient expression of miR-17 partly inhibited the hematopoietic supporting ability of FBMOB-hTERT. We further identified that HIF-1α is responsible for, at least in part, the promoted hematopoietic supporting ability of FBMOB-hTERT caused by miR-17. HIF-1α expression is markedly enhanced in miR-17 overexpressed FBMOB-hTERT upon interaction with CB CD34⁺ cells compared to other niche associated factors. More interestingly, the specific erythroid lineage expansion of CB CD34⁺ cells caused by osteoblastic miR-17 was abrogated by HIF-1α knock down.

Conclusion/Significance

Our data demonstrated that CB CD34⁺ cell expansion can be partly promoted by osteoblastic miR-17, and in particular, ectopic miR-17 can cause a specific expansion of the erythroid lineage through augmenting HIF-1α in osteoblasts.     

In Situ OH Generation from O2 ? and H2O2 Plays a Critical Role in Plasma-Induced Cell Death

Reactive oxygen and nitrogen species produced by cold atmospheric plasma (CAP) are considered to be the most important species for biomedical applications, including cancer treatment. However, it is not known which species exert the greatest biological effects, and the nature of their interactions with tumor cells remains ill-defined. These questions were addressed in the present study by exposing human mesenchymal stromal and LP-1 cells to reactive oxygen and nitrogen species produced by CAP and evaluating cell viability. Superoxide anion (O₂ ⁻) and hydrogen peroxide (H₂O₂) were the two major species present in plasma, but their respective concentrations were not sufficient to cause cell death when used in isolation; however, in the presence of iron, both species enhanced the cell death-inducing effects of plasma. We propose that iron containing proteins in cells catalyze O₂ ⁻ and H₂O₂ into the highly reactive OH radical that can induce cell death. The results demonstrate how reactive species are transferred to liquid and converted into the OH radical to mediate cytotoxicity and provide mechanistic insight into the molecular mechanisms underlying tumor cell death by plasma treatment.     

SDF2L1, a Component of the Endoplasmic Reticulum Chaperone Complex, Differentially Interacts with ��-, ��-, and ��-Defensin Propeptides

Mammalian defensins are cationic antimicrobial peptides that play a central role in host innate immunity and as regulators of acquired immunity. In animals, three structural defensin subfamilies, designated as α, β, and θ, have been characterized, each possessing a distinctive tridisulfide motif. Mature α- and β-defensins are produced by simple proteolytic processing of their prepropeptide precursors. In contrast, the macrocyclic θ-defensins are formed by the head-to-tail splicing of nonapeptides excised from a pair of prepropeptide precursors. Thus, elucidation of the θ-defensin biosynthetic pathway provides an opportunity to identify novel factors involved in this unique process. We incorporated the θ-defensin precursor, proRTD1a, into a bait construct for a yeast two-hybrid screen that identified rhesus macaque stromal cell-derived factor 2-like protein 1 (SDF2L1), as an interactor. SDF2L1 is a component of the endoplasmic reticulum (ER) chaperone complex, which we found to also interact with α- and β-defensins. However, analysis of the SDF2L1 domain requirements for binding of representative α-, β-, and θ-defensins revealed that α- and β-defensins bind SDF2L1 similarly, but differently from the interactions that mediate binding of SDF2L1 to pro-θ-defensins. Thus, SDF2L1 is a factor involved in processing and/or sorting of all three defensin subfamilies.Mammalian defensins are tridisulfide-containing antimicrobial peptides that contribute to innate immunity in all species studied to date. Defensins are comprised of three structural subfamilies: the α-, β-, and θ-defensins (1). α- and β-Defensins are peptides of about 29–45-amino acid residues with similar three-dimensional structures. Despite their similar tertiary conformations, the disulfide motifs of α- and β-defensins differ. Expression of human α-defensins is tissue-specific. Four myeloid α-defensins (HNP1–4) are expressed predominantly by neutrophils and monocytes wherein they are packaged in granules, while two enteric α-defensins (HD-5 and HD-6) are expressed at high levels in Paneth cells of the small intestine. Myeloid α-defensins constitute about 5% of the protein mass of human neutrophils. HNPs are discharged into the phagosome during phagocytic ingestion of microbial particles. HD-5 and HD-6 are produced and stored as propeptides in Paneth cell granules and are processed extracellularly by intestinal trypsin (2). β-Defensins are produced primarily by various epithelia (e.g. skin, urogenital tract, airway) and are secreted by the producing cells in their mature forms. In contrast to pro-α-defensins, which contain a conserved prosegment of ∼40 amino acids, the prosegments in β-defensins vary in length and sequence. θ-Defensins are found only in Old World monkeys and orangutans and are the only known circular peptides in animals. These 18-residue macrocyclic peptides are formed by ligation of two nonamer sequences excised from two precursor polypeptides, which are truncated versions of ancestral α-defensins. Like myeloid α-defensins, θ-defensins are stored primarily in neutrophil and monocyte granules (3).Numerous laboratories have demonstrated that the antimicrobial properties of defensins derive from their ability to bind and disrupt target cell membranes (4), and studies have shown defensins to be active against Gram-positive and -negative bacteria (5), viruses (6–9), fungi (10, 11), and parasites such as Giardia lamblia (12). Defensins also play a regulatory role in acquired immunity as they are known to chemoattract T lymphocytes, monocytes, and immature dendritic cells (13, 14), act as adjuvants, stimulate B cell responses, and up-regulate proliferation and cytokine production by spleen cells and T helper cells (15, 16).Defensins are produced as pre-propeptides and undergo post-translational processing to form the mature peptides. While much has been learned about regulation of defensin expression, little is known about the factors involved in their biosynthesis. Valore and Ganz (17) investigated the processing of defensins in cultured cells and demonstrated that maturation of HNPs occurs through two proteolytic steps that lead to formation of mature α-defensins, but the proteases involved have yet to be identified. Moreover, there are virtually no published data regarding endoplasmic reticulum (ER)² factors that are responsible for the folding, processing, and sorting steps necessary for defensin maturation and secretion or trafficking to the proper subcellular compartment. It is likely that several chaperones, proteases, and protein-disulfide isomerase (PDI) family proteins are involved. Consistent with this possibility, Gruber et al. (18) recently demonstrated the role of a PDI in biosynthesis of cyclotides, small ∼30-residue macrocyclic peptides produced by plants.The primary structures of α- and θ-defensin precursors are closely related. We therefore undertook studies to identify proteins that interact with representative propeptides of each defensin subfamily with the goal of determining common and unique processes that regulate biosynthesis of α- and θ-defensins. We used two-hybrid analysis to first identify interactors of the θ-defensin precursor, proRTD1a. As described, we identified SDF2L1, a component of the ER-chaperone complex as an interactor, and showed that it also specifically interacts with α- and β-defensins. This suggests that SDF2L1 is involved in the maturation/trafficking of defensins at a step common to all three subfamilies of mammalian defensins.     

Identifying the African Wintering Grounds of Hybrid Flycatchers Using a Multi–Isotope (δ 2H, δ 13C, δ 15N) Assignment Approach

Migratory routes and wintering grounds can have important fitness consequences, which can lead to divergent selection on populations or taxa differing in their migratory itinerary. Collared (Ficedula albicollis) and pied (F. hypoleuca) flycatchers breeding in Europe and wintering in different sub-Saharan regions have distinct migratory routes on the eastern and western sides of the Sahara desert, respectively. In an earlier paper, we showed that hybrids of the two species did not incur reduced winter survival, which would be expected if their migration strategy had been a mix of the parent species'' strategies potentially resulting in an intermediate route crossing the Sahara desert to different wintering grounds. Previously, we compared isotope ratios and found no significant difference in stable-nitrogen isotope ratios (δ ¹⁵N) in winter-grown feathers between the parental species and hybrids, but stable-carbon isotope ratios (δ ¹³C) in hybrids significantly clustered only with those of pied flycatchers. We followed up on these findings and additionally analyzed the same feathers for stable-hydrogen isotope ratios (δ ²H) and conducted spatially explicit multi-isotope assignment analyses. The assignment results overlapped with presumed wintering ranges of the two species, highlighting the efficacy of the method. In contrast to earlier findings, hybrids clustered with both parental species, though most strongly with pied flycatcher.     

Smad4 Loss Synergizes with TGFα Overexpression in Promoting Pancreatic Metaplasia,PanIN Development,and Fibrosis

Aims

While overexpression of TGFα has been reported in human pancreatic ductal adenocarcinoma (PDAC), mice with overexpressed TGFα develop premalignant pancreatic acinar-to-ductal metaplasia (ADM) but not PDAC. TGF-β signaling pathway is pivotal to the development of PDAC and tissue fibrosis. Here we sought to investigate the interplay between TGFα and TGF-β signaling in pancreatic tumorigenesis and fibrosis, namely via Smad4 inactivation.

Methods

The MT-TGFα mouse was crossed with a new Smad4 conditional knock-out mouse (Smad4^flox/flox;p48-Cre or S4) to generate Smad4^flox/flox;MT-TGFα;p48-Cre (STP). After TGFα overexpression was induced with zinc sulfate water for eight months, the pancreata of the STP, MT-TGFα, and S4 mice were examined for tumor development and fibrotic responses. PanIN lesions and number of ducts were counted, and proliferation was measured by Ki67 immunohistochemistry (IHC). Qualitative analysis of fibrosis was analyzed by Trichrome Masson and Sirius Red staining, while vimentin was used for quantification. Expression analyses of fibrosis, pancreatitis, or desmoplasia associated markers (α-SMA, Shh, COX-2, Muc6, Col1a1, and Ctgf) were performed by IHC and/or qRT-PCR.

Results

Our STP mice exhibited advanced ADM, increased fibrosis, increased numbers of PanIN lesions, overexpression of chronic pancreatitis-related marker Muc6, and elevated expression of desmoplasia-associated marker Col1A1, compared to the MT-TGFα mice. The inactivation of Smad4 in the exocrine compartment was responsible for both the enhanced PanIN formation and fibrosis in the pancreas. The phenotype of the STP mice represents a transient state from ADMs to PanINs, closely mimicking the interface area seen in human chronic pancreatitis associated with PDAC.

Conclusion

We have documented a novel mouse model, the STP mice, which displayed histologic presentations reminiscent to those of human chronic pancreatitis with signs of early tumorigenesis. The STP mice could be a suitable animal model for interrogating the transition of chronic pancreatitis to pancreatic cancer.     

Brucella Cyclic β-1,2-Glucan Plays a Critical Role in the Induction of Splenomegaly in Mice

Brucella, the etiological agent of animal and human brucellosis, is a bacterium with the capacity to modulate the inflammatory response. Cyclic β-1,2-glucan (CβG) is a virulence factor key for the pathogenesis of Brucella as it is involved in the intracellular life cycle of the bacteria. Using comparative studies with different CβG mutants of Brucella, cgs (CβG synthase), cgt (CβG transporter) and cgm (CβG modifier), we have identified different roles for this polysaccharide in Brucella. While anionic CβG is required for bacterial growth in low osmolarity conditions, the sole requirement for a successful Brucella interaction with mammalian host is its transport to periplasmic space. Our results uncover a new role for CβG in promoting splenomegaly in mice. We showed that CβG-dependent spleen inflammation is the consequence of massive cell recruitment (monocytes, dendritics cells and neutrophils) due to the induction of pro-inflammatory cytokines such as IL-12 and TNF-α and also that the reduced splenomegaly response observed with the cgs mutant is not the consequence of changes in expression levels of the characterized Brucella PAMPs LPS, flagellin or OMP16/19. Complementation of cgs mutant with purified CβG increased significantly spleen inflammation response suggesting a direct role for this polysaccharide.     

Lactobacillus reuteri Prevents Diet-Induced Obesity,but not Atherosclerosis,in a Strain Dependent Fashion in Apoe?/? Mice

Objective

To investigate whether the specific strains of Lactobacillus reuteri modulates the metabolic syndrome in Apoe−/− mice.

Methods

8 week-old Apoe−/− mice were subdivided into four groups who received either L. reuteri ATCC PTA 4659 (ATCC), DSM 17938 (DSM), L6798, or no bacterial supplement in the drinking water for 12 weeks. The mice were fed a high-fat Western diet with 0.2% cholesterol and body weights were monitored weekly. At the end of the study, oral glucose and insulin tolerance tests were conducted. In addition, adipose and liver weights were recorded along with analyses of mRNA expression of ileal Angiopoietin-like protein 4 (Angptl4), the macrophage marker F4/80 encoded by the gene Emr1 and liver Acetyl-CoA carboxylase 1 (Acc1), Fatty acid synthase (Fas) and Carnitine palmitoyltransferase 1a (Cpt1a). Atherosclerosis was assessed in the aortic root region of the heart.

Results and Conclusions

Mice receiving L. reuteri ATCC gained significantly less body weight than the control mice, whereas the L6798 mice gained significantly more. Adipose and liver weights were also reduced in the ATCC group. Serum insulin levels were lower in the ATCC group, but no significant effects were observed in the glucose or insulin tolerance tests. Lipogenic genes in the liver were not altered by any of the bacterial treatments, however, increased expression of Cpt1a was found in the ATCC group, indicating increased β-oxidation. Correspondingly, the liver trended towards having lower fat content. There were no effects on inflammatory markers, blood cholesterol or atherosclerosis. In conclusion, the probiotic L. reuteri strain ATCC PTA 4659 partly prevented diet-induced obesity, possibly via a previously unknown mechanism of inducing liver expression of Cpt1a.     

Prolonged fasting activates hypoxia inducible factors-1α, -2α and -3α in a tissue-specific manner in northern elephant seal pups

Hypoxia inducible factors (HIFs) are important regulators of energy homeostasis and cellular adaptation to low oxygen conditions. Northern elephant seals are naturally adapted to prolonged periods (1–2 months) of food deprivation (fasting) which result in metabolic changes that may activate HIF-1. However, the effects of prolonged fasting on HIFs are not well defined. We obtained the full-length cDNAs of HIF-1α and HIF-2α, and partial cDNA of HIF-3α in northern elephant seal pups. We also measured mRNA and nuclear protein content of HIF-1α, -2α, -3α in muscle and adipose during prolonged fasting (1, 3, 5 & 7 weeks), along with mRNA expression of HIF-mediated genes, LDH and VEGF. HIF-1α, -2α and -3α are 2595, 2852 and 1842 bp and encode proteins of 823, 864 and 586 amino acid residues with conserved domains needed for their function (bHLH and PAS) and regulation (ODD and TAD). HIF-1α and -2α mRNA expression increased 3- to 5-fold after 7 weeks of fasting in adipose and muscle, whereas HIF-3α increased 5-fold after 7 weeks of fasting in adipose. HIF-2α protein expression was detected in nuclear fractions from adipose and muscle, increasing approximately 2-fold, respectively with fasting. Expression of VEGF increased 3-fold after 7 weeks in adipose and muscle, whereas LDH mRNA expression increased 12-fold after 7 weeks in adipose. While the 3 HIFα genes are expressed in muscle and adipose, only HIF-2α protein was detectable in the nucleus suggesting that HIF-2α may contribute more significantly in the up-regulation of genes involved in the metabolic adaptation during fasting in the elephant seal.     

Agistemus aimogastaensis sp. n. (Acari,Actinedida, Stigmaeidae), a recently discovered predator of eriophyid mites Aceria oleae and Oxycenus maxwelli,in?olive orchards in Argentina

A new species, Agistemus aimogastaensis, is described with the aid of optical and Scanning Electron Microscopy. This mite is an important predator of two eriophyid mites (Aceria oleae and Oxycenus maxwelli) in olive orchards (Olea europaea, variety Arauco) in La Rioja Province. The problems related to eriophyids in olive orchards in Argentina are highlighted and photos of the damage on leaves and fruit are included.     

Telomerase-associated Protein 1, HSP90, and Topoisomerase II�� Associate Directly with the BLM Helicase in Immortalized Cells Using ALT and Modulate Its Helicase Activity Using Telomeric DNA Substrates

The BLM helicase associates with the telomere structural proteins TRF1 and TRF2 in immortalized cells using the alternative lengthening of telomere (ALT) pathways. This work focuses on identifying protein partners of BLM in cells using ALT. Mass spectrometry and immunoprecipitation techniques have identified three proteins that bind directly to BLM and TRF2 in ALT cells: telomerase-associated protein 1 (TEP1), heat shock protein 90 (HSP90), and topoisomerase IIα (TOPOIIα). BLM predominantly co-localizes with these proteins in foci actively synthesizing DNA during late S and G₂/M phases of the cell cycle when ALT is thought to occur. Immunoprecipitation studies also indicate that only HSP90 and TOPOIIα are components of a specific complex containing BLM, TRF1, and TRF2 but that this complex does not include TEP1. TEP1, TOPOIIα, and HSP90 interact directly with BLM in vitro and modulate its helicase activity on telomere-like DNA substrates but not on non-telomeric substrates. Initial studies suggest that knockdown of BLM in ALT cells reduces average telomere length but does not do so in cells using telomerase.Bloom syndrome (BS)⁴ is a genetic disease caused by mutation of both copies of the human BLM gene. It is characterized by sun sensitivity, small stature, immunodeficiency, male infertility, and an increased susceptibility to cancer of all sites and types. The high incidence of spontaneous chromosome breakage and other unique chromosomal anomalies in cells from BS patients indicate an increase in homologous recombination in somatic cells (1). Another notable feature of non-immortalized and immortalized cells from BS individuals is the presence of telomeric associations (TAs) between homologous chromosomes (2). Work from our group and others have suggested a role for BLM in recombination-mediated mechanisms of telomere elongation or ALT (alternative lengthening of telomeres), processes that maintain/elongate telomeres in the absence of telomerase (3–5). However, the exact mechanism by which BLM contributes to telomere stability is unknown.Several proteins interact with and regulate BLM helicase activity, including two telomere-specific proteins, TRF1 and TRF2 (6, 7). Although TRF2 stimulates BLM unwinding of telomeric and non-telomeric 3′-overhang substrates, TRF1 inhibits BLM unwinding of telomeric substrates. TRF2-mediated stimulation of BLM helicase activity on a telomeric substrate is observed when TRF2 is present in excess or with equimolar amount of TRF1 but not when TRF1 is present in molar excess. Both proteins associate with BLM specifically in ALT cells in vivo, suggesting their involvement in the ALT pathways. In addition to TRF1 and TRF2, the telomere single-strand DNA-binding protein POT1 strongly stimulates BLM helicase activity on long telomeric forked duplexes and D-loop structures (8). Other proteins also play an important role in telomere maintenance in telomerase-negative cells, including RAD50, NBS1, and MRE11, which co-localize with TRF1 and TRF2 in specialized ALT-associated promyelocytic leukemia (PML) nuclear bodies (APBs) (9–11). Thus, we hypothesize that BLM complex formation may be essential for the ALT mechanism, and its modification may occur dynamically during the specific nucleic acid transactions required to protect the telomere in cells using the ALT pathways.This study has identified previously unknown protein partners of BLM and TRF2 in ALT cells using double immunoprecipitation and mass spectrometry (MS). These include telomerase-associated protein 1 (TEP1), heat shock protein 90 (HSP90), and topoisomerase IIα (TOPOIIα). These proteins associate with BLM and TRF2 in cells using ALT but not in cells using telomerase and directly interact with BLM in vitro. This complex of proteins localizes to sites of new DNA synthesis in vivo in ALT cells, suggesting a role in telomere maintenance. We also identified HSP90 and TOPOIIα in another ALT-specific complex consisting of BLM, TRF1, and TRF2 but not TEP1. In vitro analyses demonstrate that HSP90 inhibits BLM helicase activity using both telomeric and non-telomeric substrates, whereas TEP1 and TOPOIIα initially slow the kinetics of BLM unwinding only using telomeric substrates. These findings suggest the presence of dynamic BLM-associated ALT complexes that include previously unidentified interacting proteins. The function of TEP1 in the BLM·TRF2 complex remains unclear, although its previously described interaction with the RNA subunit of telomerase (12) suggests an interesting hypothesis of cross-talk between mechanisms of telomere elongation.     

Angiostrongylus cantonensis infection in molluscs in the municipality of S?o Gon?alo,a metropolitan area of Rio de Janeiro,Brazil: role of the invasive species Achatina fulica in parasite transmission dynamics

The aim of this study was to analyse the infection dynamics ofAngiostrongylus cantonensis in its possible intermediate hosts over two years in an urban area in the state of Rio de Janeiro where the presence ofA. cantonensis had been previously recorded in molluscs. Four of the seven mollusc species found in the study were exotic.Bradybaena similaris was the most abundant, followed byAchatina fulica, Streptaxis sp., Subulina octona, Bulimulus tenuissimus, Sarasinula linguaeformis and Leptinaria unilamellata. Only A. fulica and B. similaris were parasitised by A. cantonensis and both presented co-infection with other helminths. The prevalence of A. cantonensisin A. fulica was more than 50% throughout the study. There was an inverse correlation between the population size ofA. fulica and the prevalence of A. cantonensis and abundance of the latter was negatively related to rainfall. The overall prevalence of A. cantonensis in B. similariswas 24.6%. A. fulica was the most important intermediary host of A. cantonensis in the studied area andB. similaris was secondary in importance for A. cantonensis transmission dynamics.     

Taxonomy of the Cryptopygus complex. I. Pauropygus -?a new worldwide littoral genus (Collembola,Isotomidae)

In this paper, we describe the new genus Pauropygus gen. n. which includes three minute species, blind and unpigmented, living in interstitial littoral habitats in tropical or subtropical countries. Two of these species are new to science (type species Pauropygus projectus sp. n. from New Caledonia and Pauropygus pacificus sp. n. from China); the third one, originally described in the genus Cryptopygus (Cryptopygus caussaneli Thibaud, 1996), has a larger pantropical distribution. We synonymize here Cryptopygus riebi Barra, 1997 from South Africa with Pauropygus caussaneli. Two paratypes of the Mexican species Cryptopygus axayacatl Palacios & Thibaud, 2001 turned also to be Pauropygus caussaneli, while the holotype and remaining paratypes of this species support its placement in Proisotomodes. Among the Cryptopygus complex, Pauropygus gen. n. is easily recognized by characters of mouthparts (presence of two large projections on pleural fold, basolateral field with 6 chaetae, modified mouthparts) and reduced sensillar chaetotaxy (tergal sensilla 2-3,0-1/0-1,0-1,1-2,1-2,1-3, microsensilla reduced in number: 00/0-100, with sensilla situated in p-row on the abdomen). Small size, absence of eyes and pigment are also shared by all its species. The three species belonging to the genus differ by sensillar chaetotaxy.