Effective combination of arugula vermicompost, chitin and inhibitory bacteria for suppression of the root-knot nematode Meloidogyne javanica and explanation of their beneficial properties based on microbial analysis.

Root-knot nematodes (Meloidogyne spp.) are dangerous parasites of many crops worldwide. The threat of chemical nematicides has led to increasing interest in studying the inhibitory effects of organic amendments and bacteria on plant-parasitic nematodes, but their combination has been less studied. One laboratory and four glasshouse experiments were conducted to study the effect on M. javanica of animal manure, common vermicompost, shrimp shells, chitosan, compost and vermicompost from castor bean, chinaberry and arugula, and the combination of arugula vermicompost with some bacteria, isolated from vermicompost or earthworms. The extract of arugula compost and vermicompost, common vermicompost and composts from castor bean and chinaberry reduced nematode egg hatch by 12-32% and caused 13-40% mortality of second-stage juveniles in vitro. Soil amendments with the combination vermicompost of arugula + Pseudomonas. resinovorans + Sphingobacterium daejeonense + chitosan significantly increased the yield of infected tomato plants and reduced nematode reproduction factor by 63.1-76.6%. Comparison of chemical properties showed that arugula vermicompost had lower pH, EC, and C/N ratio than arugula compost. Metagenomics analysis showed that Bacillus, Geodermatophilus, Thermomonas, Lewinella, Pseudolabrys and Erythrobacter were the major bacterial genera in the vermicompost of arugula. Metagenomics analysis confirmed the presence of chitinolytic, detoxifying and PGPR bacteria in the vermicompost of arugula. The combination of arugula vermicompost + chitosan + P. resinovorans + S. daejeonense could be an environmentally friendly approach to control M. javanica.

Intraspecific variations of morphometric indices of some species of the genus Ditylenchus Filipjev, 1936 (Nematoda: Anguinidae) in relation to diet and temperature.

In a study, 13 known species of Ditylenchus Filipjev, 1936 were collected during 20132017 mainly in the southern provinces of Iran, and the variability of their morphological and morphometric characteristics was investigated. The results showed that the number of lateral lines, V and V indices, stylet and spicule length, whose coefficient of variation was less than 10%, were more stable than the other characters. The effects of different temperatures and food sources, including several plants, fungi and carrot disc on the taxonomic characters of three populations of Ditylenchus destructor, two populations of D. dipsaci, and one population of D. myceliophagus were studied. The results showed that indices V, V and MB were more stable than the others. The indices were more affected by food source than by temperature. In addition, the calculation of the correlation between some morphometric indices showed that the distances between the anterior end and vulva, and the anterior end and anus were significantly correlated with body length in all species and populations studied.

Analyses of morphological and molecular characteristics of Telotylenchinae from Iran point at the validity of the genera Bitylenchus and Sauertylenchus.

To investigate relationships within the subfamily Telotylenchinae, more than 500 soil samples were collected from various natural and agricultural habitats in several localities of Iran. Individuals of seven known species, including Bitylenchus dubius, B. parvus, B. serranus, Sauertylenchus maximus, Tylenchorhynchus clarus, T. microconus and Trophurus ussuriensis, were recovered and characterized based on morphological and morphometric characters. Furthermore, phylogenetic relationships within representatives of the subfamily Telotylenchinae were explored using 13 sequences from the D2-D3 expansion regions of 28S ribosomal (r) DNA and 13 sequences for the partial 18S rDNA genes obtained in the present study. Analyses based on the relatively conserved 18S rDNA gene underlined the polyphyletic status of Tylenchorhynchus sensu lato, as three clusters representing Tylenchorhynchus, Bitylenchus and Sauertylenchus were well separated, and interspersed by, e.g., representatives of the Macrotrophurinae. Analysis of the more variable D2-D3 28S rDNA sequences suggested at least four clades within the subfamily Telotylenchinae: two clades dominated by Tylenchorhynchus species; a third clade harbouring representatives of Sauertylenchus, Bitylenchus, and Paratrophurus in distinct but unresolved branches;and a fourth clade dominated by Trophurus species. Although no molecular information was available from a number of genera that were originally gathered under Tylenchorhynchus sensu lato, we concluded that both Bitylenchus and Sauertylenchus should be seen as valid Telotylenchinae genera related to, but distinct from Tylenchorhynchus.

Biocontrol Potential of the Vorticella sp. Isolated from Vermicompost Against Meloidogyne javanica.

Non-chemical control of root-knot nematodes (Meloidogyne spp.), the most important plant-parasitic nematodes, is the objective of many studies. In this study, the biocontrol potential of a protozoan Vorticella sp., isolated from vermicompost, against the root-knot nematode M. javanica was investigated. The effect of different vermicompost extracts (VE) with and without Vorticella sp. on M. javanica was investigated in vitro and in glasshouse experiments on tomato plants. The result showed that Vorticella sp. + VE had an inhibitory effect on egg hatching in vitro, which was reduced by 43-62%. In addition, the results of glasshouse experiments showed that Vorticella sp. + VE significantly increased the shoot weight of infected tomato plants compared to the control and reduced the reproduction factor of M. javanica by 74-91%. Vorticella sp. has inhibitory properties against root-knot nematode and its biocontrol potential was demonstrated in this study.

Effects of Arugula Vermicompost on the Root-Knot Nematode (Meloidogyne javanica) and the Promotion of Resistance Genes in Tomato Plants.

Root-knot nematodes are the most important plantparasitic nematodes worldwide. Many efforts have been made to find non-chemical, risk-free, and environmentally friendly methods for nematode control. In this study, the effects of compost and vermicompost of arugula (Eruca sativa) on Meloidogyne javanica were investigated in three glasshouse experiments. In addition, the expression of the defense-related genes nonexpressor of pathogenesis-related 1 (NPR1) and lipoxygenase 1 (LOX1) was detected in tomato plants treated with vermicompost of arugula at 0, 2, 7, and 14 days after nematode inoculation. The result showed that the vermicompost of arugula significantly reduced the reproduction factor of the nematode by 54.4% to 70.5% in the three experiments and increased the dry weight of shoots of infected tomato plants. Gene expression analysis showed that LOX1 expression increased on the second and seventh day after nematode inoculation, while NPR1 expression decreased. The vermicompost of arugula showed stronger nematode inhibitory potential than the vermicompost of animal manure. The vermicompost of arugula is superior to arugula compost in suppressing the activity of M. javaniva and reducing its impact. It manipulates the expression of resistance genes and could induce systemic resistance against rootknot nematodes.

Molecular phylogeny of Diphtherophora de Man, 1880 (Nematoda: Diphtherophoridae), with description of a new species.

The poorly-known nematodes of the genus Diphtherophora are soil inhabitants often found in the vicinity of plant roots. In the present study, we characterize a new species belonging to the genus Diphtherophora, named D. eldarica n. sp., from Iran using both morphological and molecular approaches. The new species is characterized by striated cuticle at the anterior end of the body, head offset from the body contour, spear 15-17 µm in length, rod-shaped sperm, overhanging cuticle on the anterior lip of the anus, and a conical tail bent dorsally near the terminus. Diphterophora eldarica n. sp. also lacks ventromedian neck papillae whereas male specimens bear two precloacal ventromedian supplements at the level of the spicules. Additionally, we provide morphological and molecular data for four known Diphtherophora species including D. geraerti, D. caudata, D. perplexans, and D. tenera collected from soils of different plants and localities in Iran. Using the D2-D3 expansion segments of the 28S ribosomal (rRNA) gene including D. eldarica n. sp. and additional known species from Iran, we explore for the first time species relationships in the genus Diphtherophora within a molecular phylogenetic framework. Our results support: 1) the monophyly of the genus Diphtherophora with respect to the outgroup taxa (Tylolaimophorus and representatives of Trichodoridae), 2) the existence of six strongly supported clades within Diphtherophora, and 3) a sister relationship between D. eldarica n. sp. and a clade formed by D. perplexans and Diphtherophora sp. from the USA. Finally, this study emphasizes the diversity of the genus Diphtherophora in Iran.

Description and molecular phylogeny of Mesocriconema abolafiai n. sp. (Nematoda: Criconematidae) from Iran.

Mesocriconema abolafiai n. sp. is described by morphological, morphometric, and molecular approaches. The new species is characterized by a body slightly curved with 402 to 612 µm length, 90 to 113 cuticular body annuli with smooth to irregular margins lacking of crenation with not more than one anastomoses, lip region not offset, small flattened submedian lobes, stylet robust (52.8-60.0 µm) with well-developed knobs, open vulva with simple anterior lip, straight vagina, filled spermatheca with globular sperms, presence of males, and conical-acute tail with last annulus bilobed or rounded. Discussions are made on the characterization of M. abolafiai n. sp. from the most closely related species, M. ozarkiense, and several other species having similar tail shape. Furthermore, results of phylogenetic analyses inferred from D2 to D3 expansion fragments of 28S rRNA, 18S rRNA, and ITS rRNA gene sequences revealed the phylogenetic position of the new species within representatives of Criconematidae and supported morphological justifications for considering this population from Iran as a new species in the genus Mesocriconema.Mesocriconema abolafiai n. sp. is described by morphological, morphometric, and molecular approaches. The new species is characterized by a body slightly curved with 402 to 612 µm length, 90 to 113 cuticular body annuli with smooth to irregular margins lacking of crenation with not more than one anastomoses, lip region not offset, small flattened submedian lobes, stylet robust (52.8-60.0 µm) with well-developed knobs, open vulva with simple anterior lip, straight vagina, filled spermatheca with globular sperms, presence of males, and conical-acute tail with last annulus bilobed or rounded. Discussions are made on the characterization of M. abolafiai n. sp. from the most closely related species, M. ozarkiense, and several other species having similar tail shape. Furthermore, results of phylogenetic analyses inferred from D2 to D3 expansion fragments of 28S rRNA, 18S rRNA, and ITS rRNA gene sequences revealed the phylogenetic position of the new species within representatives of Criconematidae and supported morphological justifications for considering this population from Iran as a new species in the genus Mesocriconema.

New and known species of Nothotylenchus Thorne, 1941 (Nematoda: Anguinidae) from Iran with an updated list of species.

During a survey conducted during 2013-2017, five known and two new species of Nothotylenchus Thorne, 1941 were collected from the southern provinces of Iran. N. brzeskii n. sp. is characterised by a body length of 774-922 µm, lateral fields with four incisures, delicate, short stylet (7-8 µm) with small rounded knobs, pyriform, offset or slightly overlapping basal pharyngeal bulb, posterior vulva position (V = 83.4-84.4), short PUS (5-10 µm), spicules 20.5-23 µm long, and thick tail with rounded to dull terminus. N. siddiqi n. sp. is characterised by a body length of 573-645 µm, six to nine incisures in lateral fields, delicate, short stylet (6.5-7.5 µm) with rounded knobs, pyriform or slightly elongate and offset basal pharyngeal bulb, V = 79.3-81.0, PUS = 26.5-40 µm, short spicules = 14.5-16.5 µm, and tail with rounded terminus. Morphometric data of the studied species are presented and intraspecific variation of their morphometrics and morphological characters is discussed. The list of world Nothotylenchus species is updated, and a dichotomous identification key and an updated tabular compendium for 41 valid species are provided.

Genetic intraspecific diversity of Meloidogyne javanica parasitizing vegetables in southern Iran.

In order to investigate different species of root-knot nematodes associated with vegetable production in southern regions of Iran, 37 populations of the most predominant species, Meloidogyne javanica, were recovered. Morphological and morphometric studies were carried on the characters of females, males, J2s and perineal patterns. Species-specific Sequence Characterized Amplified Region (SCAR) primers confirmed morphological studies, and all these populations produced specific band in 670 bp using Fjav and Rjav primers. Genetic diversity of different populations was studied by Inter Simple Sequence Repeats (ISSR) and Random Amplified Polymorphic DNA (RAPD) markers implementing 10 primers for each approach. Results revealed a relatively low genetic diversity (the percentage of polymorphic bands were 19.08 and 24.60 for ISSR and RAPD, respectively). The analyses of molecular variance indicated that the variation resulted from genotypic variations within region and variances among regions are 81% and 19% for RAPD, and 86% and 14% for ISSR, respectively. On the other hand, F ST and Nm values are 0.140 and 1.535 for ISSR while these values are 0.188 and 1.079 for RAPD. So it can be concluded that there is a great deal of gene flow between populations due to the movement of plant material contaminated with nematodes, which results in high mixing between populations. ISSR and RAPD datasets failed to group populations according to their geographic region. There were no pathotypes or other intraspecific biological entities observed in the species.In order to investigate different species of root-knot nematodes associated with vegetable production in southern regions of Iran, 37 populations of the most predominant species, Meloidogyne javanica, were recovered. Morphological and morphometric studies were carried on the characters of females, males, J2s and perineal patterns. Species-specific Sequence Characterized Amplified Region (SCAR) primers confirmed morphological studies, and all these populations produced specific band in 670 bp using Fjav and Rjav primers. Genetic diversity of different populations was studied by Inter Simple Sequence Repeats (ISSR) and Random Amplified Polymorphic DNA (RAPD) markers implementing 10 primers for each approach. Results revealed a relatively low genetic diversity (the percentage of polymorphic bands were 19.08 and 24.60 for ISSR and RAPD, respectively). The analyses of molecular variance indicated that the variation resulted from genotypic variations within region and variances among regions are 81% and 19% for RAPD, and 86% and 14% for ISSR, respectively. On the other hand, F ST and Nm values are 0.140 and 1.535 for ISSR while these values are 0.188 and 1.079 for RAPD. So it can be concluded that there is a great deal of gene flow between populations due to the movement of plant material contaminated with nematodes, which results in high mixing between populations. ISSR and RAPD datasets failed to group populations according to their geographic region. There were no pathotypes or other intraspecific biological entities observed in the species.

Early transcriptional responses to soybean cyst nematode HG Type 0 show genetic differences among resistant and susceptible soybeans.

KEY MESSAGE: Root transcriptome profiling of three soybean cultivars and a wild relative infected with soybean cyst nematode at migratory phase revealed differential resistance pathway responses between resistant and susceptible genotypes. The soybean cyst nematode (SCN), Heterodera glycines, is the most serious pathogen of soybean production throughout the world. Using resistant cultivars is the primary management strategy against SCN infestation. To gain insight into the still obscure mechanisms of genetic resistance to nematodes in different soybean genotypes, RNA-Seq profiling of the roots of Glycine max cv. Peking, Fayette, Williams 82, and a wild relative (Glycine soja PI 468916) was performed during SCN infection at the migratory phase. The analysis showed statistically significant changes of expression beginning at eight hours after inoculation in genes associated with defense mechanisms and pathways, such as the phenylpropanoid biosynthesis pathway, plant innate immunity and hormone signaling. Our results indicate the importance of the early plant response to migratory phase nematodes in pathogenicity determination. The transcriptome changes occurring during early SCN infection included a number of genes and pathways specific to the different resistant genotypes. We observed the most extensive resistant transcriptome reaction in PI 468916, where the resistant response was qualitatively different from that of commonly used G. max varieties.

Description of Ditylenchus paraparvus n. sp. from Iran with an updated list of Ditylenchus Filipjev, 1936 (Nematoda: Anguinidae).

During a survey, 12 known and one new species of Ditylenchus Filipjev, 1936 were collected from southern provinces of Iran during 2013-2017. Ditylenchus paraparvus n. sp. is characterised by a short body length (441-543 µm), lateral field with four lines, delicate and short stylet (6-7.5 µm) with small rounded or posteriorly sloping knobs, pyriform and offset basal pharyngeal bulb, V = 67.7-76.4, short post-vulval uterine sac less than one vulval body width long, small spicules (12.5-15 µm) and almost cylindrical tail (71-90 µm) with rounded end. Morphometric data of studied species are presented and intraspecific variation of their morphometrics and morphological characters is discussed. The list of the world Ditylenchus species is updated, and a dichotomous identification key and an updated tabular compendium for 63 valid species is provided.

An updated and annotated checklist of the Tylenchulidae (Nematoda: Criconematoidea) of Iran.

The list of plant parasitic nematodes of the family Tylenchulidae, collected from Iran, is updated. Forty-one species belonging to five genera and three subfamilies are included in the list. Data for seven species of Paratylenchus are added, of which two species-P. perminimus and P. salubris-are new records for the Iranian nematofauna. The list of species, further information on their morphometrics, references to full or partial descriptions, associated plants, geographical distribution and some taxonomic remarks are provided. More detailed studies on some doubtful populations are added. Information on the taxonomic position of species in different classification schemes, as well as the preferences of some species for certain climatic conditions or ecological niches is provided.

Disruption of the Pathogenicity and Sex Ratio of the Beet Cyst Nematode Heterodera schachtii by Host-Delivered RNA Interference.

The beet cyst nematode (BCN) Heterodera schachtii causes serious damage and yield losses in numerous important crops worldwide. This study examines the efficacy of three types of transgenic Arabidopsis RNA interference (RNAi) lines to decrease the biological activity of this devastating nematode. The first RNAi construct (E1E2-RNAi) targets two nematode endoglucanase genes, which are involved in BCN pathogenicity, the second construct (MSP-RNAi) contains a fragment corresponding to the major sperm protein transcript necessary for BCN development and reproduction, and the third construct (E1E2MSP-RNAi) comprises all three target fragments. Transcript expression profiles of the target genes in all biological stages of the nematode were determined for the initial inoculated population and the resulting progeny. Bioassay data under indoor aseptic cultivation indicated that feeding on these RNAi lines did not affect pathogenic activity and reproductive capacity of the initial population, whereas inoculating the progeny into new transgenic plants corresponding with the lines from which they were recovered reduced the nematode penetration and the number of eggs per cyst. In addition, the male/female ratio increased more than the double, and the effects of RNAi continued in the second generation of the nematodes, because the progeny derived from E1E2-RNAi and E1E2MSP-RNAi lines showed an impaired ability to infect wild-type plants.

Morphological characterization of Trichotylenchus gorganiensis n. sp. (Nematoda: Dolichodoridae) and photomicrographs of several other species of the subfamily Telotylenchinae Siddiqi, 1960.

Trichotylenchus gorganiensis n. sp. is described and illustrated based on morphological and morphometric data. The new species is characterized by its 760-1073 µm long body, conoid-rounded lip region continuous with the body contour and bearing 5-7 fine striae, 22.0-24.5 µm long stylet, basal pharyngeal bulb offset or slightly overlapping intestine, post-anal sac extending 50-73 % of the tail region, and cylindrical or subclavate tail with a striated terminus. Differences of the new species from the closely related species T. astriatus, T. astriatoides, T. changlingensis and T. papyrus are discussed. Photomicrographs and several taxonomic notes on 13 other species of Telotylenchinae, collected from Iran, are provided.

Disparate gain and loss of parasitic abilities among nematode lineages.

Plant parasitism has arisen time and again in multiple phyla, including bacteria, fungi, insects and nematodes. In most of these organismal groups, the overwhelming diversity hampers a robust reconstruction of the origins and diversification patterns of this trophic lifestyle. Being a moderately diversified phylum with ≈ 4,100 plant parasites (15% of total biodiversity) subdivided over four independent lineages, nematodes constitute a major organismal group for which the genesis of plant parasitism could be mapped. Since substantial crop losses worldwide have been attributed to less than 1% of these plant parasites, research efforts are severely biased towards this minority. With the first molecular characterisation of numerous basal and supposedly harmless plant parasites as well as their non-parasitic relatives, we were able to generate a comprehensive molecular framework that allows for the reconstruction of trophic diversification for a complete phylum. In each lineage plant parasites reside in a single taxonomic grouping (family or order), and by taking the coverage of the next lower taxonomic level as a measure for representation, 50, 67, 100 and 85% of the known diversity was included. We revealed distinct gain and loss patterns with regard to plant parasitism per se as well as host exploitation strategies between these lineages. Our map of parasitic nematode biodiversity also revealed an unanticipated time reversal in which the two most ancient lineages showed the lowest level of ecological diversification and vice versa.

Numerical Taxonomy Helps Identification of Merliniidae and Telotylenchidae (Nematoda: Tylenchoidea) from Iran.

Numerical taxonomy was used for identification and grouping of the genera, species, and populations in the families Merliniidae and Telotylenchidae. The variability of each of 44 morphometric characters was evaluated by calculation of the coefficient of variability (CV) and the ratio of extremes (max/min) in the range of 1,020 measured females. Also correlation and regression analyses were made between characters to find potential collinearities. Hierarchical cluster analysis (HCA) was used for (i) grouping 21 genera in the superfamily Dolichodoroidea based on literature data coded for states of 18 diagnostic characters, and (ii) for grouping Iranian populations belonging to selected genera. Furthermore, STEPDISC analysis was used for (i) grouping 11 genera of Merliniidae and Telotylenchidae based on the measurements of 35 characters from 1,007 Iranian female specimens, and (ii) grouping measured females of eight species of Amplimerlinius and Pratylenchoides. The multivariate data analysis approach showed robust enough to summarize relationship between morphometric characters and group genera, species, and populations of the nematodes and in particular help to identify the genera and species of Amplimerlinius and Pratylenchoides.

A new and four known species of Diphtherophora (Nematoda: Diphtherophoridae) from Iran, with a diagnostic compendium of its species.

A new and four known species of the genus Diphtherophora-D. geraerti sp. n., D. caudata, D. obesa, D. perplexans and D. tenera-were collected and identified from different plants and localities in Iran. Morphological and morphometric characters of Diphtherophora geraerti sp. n. are compared with those of closely-related species, including D. siddiqii, D. brevicollis, D. lata, D. obesa, D. citri, D. vasilevi and D. vitoshae. Females of the new species have a head continuous with the body contour, spear 11-14 µm in length, rod-shaped spermatozoa and conical tail with bluntly rounded to hemispherical terminus. Males have two ventromedian neck papillae located 22-25 and 52-60 µm from the anterior end, and two ventromedian supplements at 11-13 and 75-80 µm anterior from cloacal aperture, respectively. All Diphtherophora species recovered as part of the present study are new records for the Iranian nematode fauna. Finally, a diagnostic compendium is given, to assist with species identification in the genus.

Morphological and molecular characterization of Pratylenchoides persicus n. sp. (Nematoda: Merliniidae) and additional data on two other species of the genus from Iran.

Some nematologists recently placed the genus Pratylenchoides, ("Lesion Nematode-like") in the family Merliniidae. To investigate Pratylenchoides species diversity and their relationships with other Merliniidae genera, specimens were collected from various habitats in the northern and northwestern provinces of Iran. The morphological and molecular study yielded three species of the genus Pratylenchoides, including P. persicus n. sp. This new species is characterized by having lip region rounded or slightly flattened anteriorly with four or five fine but distinct annuli, pharyngeal glands off-set or slightly overlapping the intestine dorsally, all three gland nuclei located anterior to the pharyngo-intestinal valve and tail cylindrical with truncate to low rounded terminus. Morphologically, P. persicus n. sp. can be distinguished from the most closely related species, P. heathi by having shorter body and stylet length in females and males, as well as a shorter tail with different terminus in females. Pratylenchoides laticauda and P. cf. nevadensis are reported from Iran for the first time. Phylogenetic analysis based on the D2/D3 region of the large subunit of ribosomal DNA revealed Pratylenchoides as a monophyletic genus, and it supports the delineation of the new species, P. persicus n. sp.

One new and three known species of Geocenamus Thorne & Malek, 1968 (Nematoda: Merliniidae) from Iran.

Soil samples collected from different plants and localities in Iran yielded one new and three known species of the genus Geocenamus with tessellated cuticle. Geocenamus conicaudatus n. sp. is characterised by a hemispherical cephalic region, set-off from the rest of body by a constriction, a slender stylet 17-20 µm long with laterally directed knobs, coarsely annulated cuticle with 32-34 longitudinal striae and a conical tail with smooth, finely rounded terminus. Morphological and molecular studies on the populations of G. paniculoides and G. tartuensis indicated that G. paniculoides may be considered as a valid species. Intraspecific variability of some characters of 30 Iranian populations of G. rugosus were studied. An identification key to 32 valid species of the genus Geocenamus is given.

Contribution to a revision of the genus Pratylenchoides Winslow, 1958 (Nematoda: Merliniidae), with redescription of P. erzurumensis Yüksel, 1977 from Iran.

Morphological and morphometric studies were conducted on the populations and paratypes of 14 species of the genus Pratylenchoides, including P. alkani, P. acuticauda, P. arenarius, P. arenicola, P. bacilisemenus, P. crenicauda, P. erzurumensis, P. heathi, P. laticauda, P. ritteri, P. rivalis, P. sheri, P. utahensis and P. variabilis. In addition, P. erzurumensis was redescribed based on a bisexual population from western Iran. The conoid head of males is an additional diagnostic character of this species distinguishing it from the closely related P. laticauda and P. camachoi. Based on detailed study of the main characters, synonymies of P. arenarius with P. bacilisemenus and P. variabilis with P. crenicauda are proposed. Synonymy of P. alkani with P. ritteri is supported also by morphological and biological evidence. Some taxonomic notes are made on some other species of the genus. Finally, a diagnostic compendium and key is provided for species identification.