Predominance of malolactic fermentation overd-glucose utilization inLactobacillus plantarum andLactobacillus curvatus wine strains

Summary Two selected wine strains of the genusLactobacillus (L. plantarum 197 andL. curvatus 783) were tested for their ability to complete malolactic fermentation (MLF) in a synthetic medium (PBM-broth) supplemented withL-malic acid (7.5–74.6 mM) andD-glucose (5.5–55 mM). The 24 directed fermentation assays, 12 for each bacterial strain, were carried out at 20°C and pH 3.5. MLF was completed (residualL-malic acid 0.2 mM) in eight days in 19 of the 24 fermentation assays, even in the presence of 74.6 mML-malic acid or 55.5 mMD-Glucose utilization was generally simultaneous to MLF but was completed (residual concentrations 0.2 mM) only in 6 of the 24 fermentation assays. These results support the use of these strains in directed MLF assays at the very differentL-malic acid andD-glucose concentrations tested.     

<Emphasis Type="Italic">Fusarium</Emphasis> species of the <Emphasis Type="Italic">Gibberella fujikuroi</Emphasis> complex and fumonisin contamination of pearl millet and corn in Georgia,USA

This study was designed to identify and compare the Fusarium species of the Gibberella fujikuroi complex on pearl millet (Pennisetum glaucum (L.) R. Br) and corn (Zea mays L.) crops grown in southern Georgia, and to determine their influence on potential fumonisin production. Pearl millet and corn samples were collected in Georgia in 1996, 1997 and 1998. Three percent of the pearl millet seeds had fungi similar to the Fusarium species of the G. fujikuroi species complex. One hundred and nineteen representative isolates visually similar to the G. fujikuroi species complex from pearl millet were paired with mating population A (Fusarium verticillioides (Sacc.) Nirenberg), mating population D (F. proliferatum (Matsushima) Nirenberg) and mating population F (F. thapsinum (Klittich, Leslie, Nelson and Marasas) tester strains. Successful crosses were obtained with 50.4%, 10.1% and 0.0% of these isolates with the A, D and F tester strains, while 39.5 of the isolates did not form perithecia with any tester strains. Two of the typical infertile isolates were characterized by DNA sequence comparisons and were identified as Fusarium pseudonygamai (Nirenberg and ODonnell), which is the first known isolation of this species in the United States. Based on the pattern of cross-compatibility, conidiogenesis, colony characteristics and media pigmentation, a majority of the infertile isolates belong to this species. Fumonisins FB₁ and FB₂ were not detected in any of the 81 pearl millet samples analyzed. The species of the G. fujikuroi species complex were dominant in corn and were isolated from 84%, 74% and 65% of the seed in 1996, 1997 and 1998, respectively. Representative species of the G. fujikuroi species complex were isolated from 1996 to 1998 Georgia corn survey (162, 104 and 111 isolates, respectively) and tested for mating compatibility. The incidence of isolates belonging to mating population A (F. verticillioides) ranged from 70.2% to 89.5%. Corn survey samples were assayed for fumonisins, and 63% to 91% of the 1996, 1997 and 1998 samples were contaminated. The total amount of fumonisins in the corn samples ranged from 0.6 to 33.3 g/g.     

Cloning and expression of <Emphasis Type="Italic">mel</Emphasis> gene from <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Previous work from our laboratory has shown that most of Bacillus thuringiensis strains possess the ability to produce melanin in the presence of l-tyrosine at elevated temperatures (42 °C). Furthermore, it was shown that the melanin produced by B. thuringiensis was synthesized by the action of tyrosinase, which catalyzed the conversion of l-tyrosine, via l-DOPA, to melanin. In this study, the tyrosinase-encoding gene (mel) from B. thuringiensis 4D11 was cloned using PCR techniques and expressed in Escherichia coli DH5 . A DNA fragment with 1179 bp which contained the intact mel gene in the recombinant plasmid pGEM1179 imparted the ability to synthesize melanin to the E. coli recipient strain. The nucleotide sequence of this DNA fragment revealed an open reading frame of 744 bp, encoding a protein of 248 amino acids. The novel mel gene from B.thuringiensis expressed in E. coli DH5 conferred UV protection on the recipient strain.     

Isolation and characterization of membrane-associated proteoglycans from normal and malignant human mammary epithelial cells

The plasma membrane-associated proteoglycans of a malignant human breast cell line (MDA-MB-231) were compared with the corresponding proteoglycans from a normal cell line (HBL-100). The labeled proteoglycans were isolated from the plasma membranes of cells grown in the presence of [³H]glucosamine and [³⁵S]Na₂SO₄ by extraction with guanidine hydrochloride and subsequently purified by DEAE-ion exchange chromatography. Their structural properties were established by treatment with nitrous acid, heparitinase and chondroitinase ABC, and by gel filtration before and after alkaline -elimination. About 18% of the proteoglycans synthesized by these cell lines were associated with the plasma membranes. The HBL plasma membranes contained 80% heparan sulfate and 20% chondroitin sulfate proteoglycans whereas MDA plasma membranes had 50% heparan sulfate and 50% chondroitin sulfate proteoglycans. The MDA plasma membrane contained two heparan sulfate proteoglycans, both having nearly the same molecular size as the two species secreted into the medium by these cells. The HBL plasma membrane also contained two hydrodynamic size heparan sulfate proteoglycans. The larger hydrodynamic size species has a slightly lower molecular size than that secreted into the medium, and the smaller hydrodynamic size species was not detectable in the medium. Even though the major chondroitin sulfate proteoglycans from MDA plasma membranes were smaller in size than those from HBL plasma membrane, a larger proportion of the glycosaminoglycan chains of the former were bigger than those from the latter.Abbreviations CHAPS 3-[(3-cholamidopropyl)dimethylammonio]propane-1-sulfonate - Di-OS 2-acetamido-2-deoxy-3-O-(-d-gluco-4-ene-pyranosyluronic acid)-d-galactose - Di-4S 2-acetamido-2-deoxy-3-O-(-d-gluco-4-ene-pyranosyluronic acid)-4-O-sulfo-d-galactose - Di-6S 2-acetamido-2-deoxy-3-O-(-d-gluco-4-ene-pyranosyluronic acid)-6-O-sulfo-d-galactose - Gdn-HCl guanidine hydrochloride - WGA wheat germ agglutinin     

Asperula visianii,nova spec., undA. staliana (Rubiaceae), Endemiten der Inseln Mitteldalmatiens

Asperula visianii Korica is described as a new stenoendemic species from the small Central Dalmatian island of Svetac (near Vis). It differs in several morphological features (which remain constant in cultivation) and in its ecology from the closely relatedA. staliana Vis., endemic on the nearby island of Bievo.

     

Analysis of the regulation of penicillin biosynthesis in Aspergillus nidulans by targeted disruption of the acvA gene

To analyse the regulation of the biosynthesis of the secondary metabolite penicillin in Aspergillus nidulans, a strain with an inactivated acvA gene produced by targeted disruption was used. acvA encodes -(l--aminoadipyl)-l-cysteinyl-d-valine synthetase (ACVS), which catalyses the first step in the penicillin biosynthetic pathway. To study the effect of the inactivated acvA gene on the expression of acvA and the second gene, ipnA, which encodes isopenicillin N synthase (IPNS), A. nidulans strain XEPD, with the acvA disruption, was crossed with strain AXB4A carrying acvA-uidA and ipnA-lacZ fusion genes. Ascospores with the predicted non-penicillin producing phenotype and a hybridization pattern indicating the presence of the disrupted acvA gene, and the fusion genes integrated in single copy at the chromosomal argB locus were identified. Both fusion genes were expressed at the same level as in the non-disrupted strain. Western blot analysis (immunoblotting) revealed that similar amounts of IPNS enzyme were present in both strains from 24 to 68 h of a fermentation run. In the acvA disrupted strain, IPNS and acyl-CoA: 6-aminopenicillanic acid acyltransferase (ACT) specific activities were detected, excluding a sequential induction mechanism of regulation of the penicillin biosynthesis gene ipnA and the third gene aat.     

Pyrolysis mass spectrometry characterisation and numerical taxonomy ofAeromonas spp.

Reference strains (2) and 29 isolates ofAeromonas spp. from clinical material and environmental specimens were characterised in traditional biochemical tests, and in pyrolysis mass spectrometry, which gives data reflecting whole-cell composition. Numerical taxonomic analyses of the data sets were compared with conventional identification at species level, and pathogenic potential, as inferred from the origin of the isolates. Clustering with conventional test reaction patterns showed, for each of the species represented, a clearly defined core group of typical isolates, surrounded by a halo of aberrant strains. One further cluster comprised strains intermediate betweenA. caviae andA. hydrophila, and one strain was grossly atypical in both analyses. Clustering from pyrolysis data corresponded less well with species identification. Broadly, the biochemical division between core and halo strains was supported in pyrolysis forA. caviae andA. sobria, but the main group ofA. hydrophila in pyrolysis comprised strains clustering in the core and halo groups of this species, and three strains intermediate betweenA. hydrophila andA. caviae in biochemical tests. Two further pyrolysis clusters comprised core and halo strains ofA. hydrophila. However, pyrolysis clustering correlated well with inferred pathogenicity, showing four clusters of probable pathogens, six clusters of probable nonpathogens, and one two member cluster of doubtful status. Most strains that clustered in the species haloes, or in species-intermediate groups in biochemical tests, were non-human isolates, or were isolated in the absence of symptomatic infection. The correlation of inferred pathogenicity with biochemical clustering was poorer than that with pyrolysis clustering.Abbreviations CTRP conventional test reaction pattern - PyMS pyrolysis mass spectrometry     

Fungi in bathwater and sludge of bathroom drainpipes

Samples of bathwater from 14 homes and 22 public bathhouses and sludge in drainpipes from 19 house-hold bathrooms were plated out onto potato dextrose agar supplemented with chloramphenicol. Several media were used to study colony morphology of the isolates and the thermotolerance and alkaline tolerance of each isolate were examined.Eleven sludge samples produced 12 isolates of Exophiala jeanselmei, 2 of E. dermatitidis and 1 of E. moniliae. Five household bathwater samples produced 2 isolates of E. jeanselmei, 4 of E. dermatitidis and 1 of E. alcalophila. One isolate of E. jeanselmei, 2 of E. dermatitidis, 3 of E. moniliae and 2 of unidentified Exophiala species were recovered from 6 samples of the bathwater dissolving Chinese medicine in the bathtubs of public bathhouses. One isolate of E. jeanselmei was recovered from the 15 samples of bathwater from public bathhouses. Bathwater and sludge in bathroom drainpipes may be an important habitat of Exophiala species.     

Diversity ofFrankia strains isolated from a single alder stand

One hundredFrankia strains isolated from variousAlnus species in a single alder stand were tested for plasmid presence. Plasmid DNA was observed in five of the frankiae strains and was analyzed. We found that plasmids with a similar molecular weight exhibited in fact minor divergences in restriction patterns. The genetic diversity among the five isolates which contained plasmids and seven isolates which contained no plasmid DNA were examined by using restriction endonucleas digestions, Southern hybridization ofnifHDK,nifAB genes, andFrankia cryptic DNA fragments determined at random. Results indicate that genomic DNA digestion patterns and Southern hybridizations to anifHDK probe were not able to discriminate between closely related frankiae. On the other hand, plasmid presence, Southern hybridization to anifAB proble or to a crypticFrankia probe allowed us to delineate groupings of these isolates.     

Isolation and chemical characterization of lipopolysaccharides from four Mycoplana species (M. bullata,M. segnis,M. ramosa and M. dimorpha)

Lipopolysaccharides (LPS), isolated from four Mycoplana species, i.e. the type strains of M. bullata, M. segnis, M. ramosa and M. dimorpha, were characterized onto their chemical composition and their respective lipid A-types. Those of M. bullata and M. segnis showed on DOC-PAGE an R-type character and had lipid A's of the Lipid A_DAG-type which exclusively contained 2,3-diamino-2,3-dideoxy-d-glucose as lipid A sugar. LPS's of M. ramosa and M. dimorpha showed, although only weakly expressed, ladder-like patterns on DOC-PAGE indicating some S-type LPS's and lipid A of the d-glucosamine type (Lipid A_GlcN). M. bullata LPS contained mannose and glucose in major amounts and additionally l-glycero-d-mannoheptose, whereas M. segnis LPS was composed of rhamnose, mannose and glucose together with both, d-glycero-d-manno- and l-glycero-d-manno-heptoses in a molar ratio of 1:2. All LPS's contained 2-keto-3-deoxy-octonic acid (Kdo), phosphate and an unidentified acidic component X. In addition to X, M. segnis LPS contained glucuronic and galacturonic acids, whereas M. ramosa LPS contained only galacturonic acid. Acetic acid hydrolysis of the LPS resulted in splitting off lipid A moieties, very rich in 3-hydroxy fatty acids, in particular in 3-OH-12:0 (in Lipid A_DAG), or in 3-OH-14:0 (in Lipid A_GlcN). Analysis of the 3-acyloxyacyl residues revealed major amounts of amide-linked 3-OH(3-OH-13:0)12:0 in lipid A of M. bullata and 3-OH(12:0)12:0 in lipid A of M. segnis. The rare 4-oxo-myristic acid (4-oxo-14:0) was observed only in M. bullata LPS, where it is ester-linked. Amide linked diesters could not be traced in M. ramosa and M. dimorpha. All four lipid A's lacked erster-bound acyloxyacyl residues.Non-standard abbreviations DAG 2,3-diamino-2,3-dideoxy-d-glucose - Kdo 2-keto-3-deoxy-octonate - LPS lipopolysaccharide - PITC phenyl isothiocyanate - NANA N-acetyl neuraminic acid     

Inhibition of some mycotoxigenic fungi by iturin A,a peptidolipid produced by Bacillus subtilis

Bacillus subtilis produces peptidolipid compounds of the iturin group that have been shown to have antifungal properties, but not all fungal species are sensitive to these compounds. In this study, the activity of iturin A, produced by B. subtilis strain B-3, was tested. Paper disks impregnated with various concentrations of iturin A were placed on agar plates seeded with conidia of toxigenic species of Fusarium, Gerlacia, Penicillium or Aspergillus. Most isolates were inhibited at iturin A concentrations as low as 4 g/disk. Penicillium italicum, P. vindicatum, A. ochraceus and A. versicolor were most strongly inhibited by the iturin whereas P. citrinum and A. parasiticus were least sensitive to iturin A.Mention of a trademark or proprietary product does not constitute a guarantee or warranty by the US Department of Agriculture and does not imply approval to the exclusion of other products that may also be suitable.     

Toxicity and moniliformin production by four recently described species of Fusarium and two uncertain taxa

Four recently described species, Fusarium nygamai, F. dlamini, F. beomiforme and F. napiforme and two uncertain taxa, F. nygamai from millet in Africa and Fusarium species from rice with Bakanae disease, were tested for toxicity and moniliformin production. Cultures grown on autoclaved corn were fed to groups of four one-day-old ducklings for 14 days. Isolates that caused the death of 3 or 4 out of 4 ducklings were considered to be toxic and analyzed for moniliformin. All 15 isolates of F. dlamini tested were nontoxic. The other taxa contained some isolates that were toxic to ducklings and produced moniliformin in corn cultures. This is the first report of moniliformin production by F. beomiforme (200–890 g/g), and F. napiforme (16–388 g/g), and by F. nygamai not obtained from millet in Africa (15–874 g/g). The highest production of moniliformin was obtained from the 19 isolates of F. nygamai from millet in Africa (4300–18200g/g) and the 15 isolates from rice with Bakanae disease (2300–19300 g/g). The taxonomic position of these two uncertain taxa should be re-evaluated.     

Synthesis ofNeoglycoproteins containing the 3,6-di-O-methyl-β-d-glucopyranosyl epitope and their use in serodiagnosis of leprosy

A stratagem for the synthesis ofneoglycoproteins suitable for the selective serodiagnosis of leprosy is described in which synthetic 3,6-di-O-methyl--d-glucopyranose, the epitope of phenolic glycolipid I fromMycobacterium leprae, was used. Condensation of 8-methoxycarbonyloctanol with the acetobromo derivative of 3,6-di-O-methylglucose gave 8-methoxycarbonyloctyl 2,4-di-O-acetyl-3,6-di-O-methyl--d-glucopyranoside in 65% yield, and with absolute stereospecificity for the anomer. The deacylated product was converted to the crystalline hydrazide and coupled to bovine gamma globulin, bovine serum albumin and poly-d-lysinevia intermediate acyl azide formation to produce the 8-carbonyloctyl 3,6-di-O-methyl--d-glucopyranosyl polypeptides. Theneoglycoproteins were highly sensitive in ELISA and emulated the specificity of the native glycolipid in analysis of sera from patients throughout the spectrum of leprosy and from different geographical regions. The 8-carbonyloctyl 3,6-di-O-methyl--d-glucopyranoside-bovine serum albumin was also synthesized and shown to have about one-half the activity of the -linkedneoglycoprotein. A different synthetic approach produced the 8-carbonyloctyl 4-O-(3,6-di-O-methyl--d-glucopyranosyl)--l-rhamnopyranoside-bovine serum albumin which was also highly sensitive and specific for the serodiagnosis of leprosy. The presence of the second sugar unit, similar to that in the native glycolipid but for the absence ofO-methyl groups, seemed to provide a probe with greater felicity for the serological detection of tuberculoid leprosy.Thus, the results indicate that highly sensitive and specific antigen probes for the serodiagnosis of leprosy can be constructed based only on the terminal one or two sugars of phenolic glycolipid I, and the synthetic approach leads to the formation of haptens with absolute stereospecificity.Nomenclature BGG bovine gamma globulin - PGL-I phenolic glycolipid I - PDL poly-d-lysine - PBS phophate-buffered saline - 3,6-Me₂-Glc-Link-BSA 8-carbonyloctyl 3,6-di-O-methyl-glucopyranoside-bovine senalbumin - 3,6-Me₂-Glc-Rha-Link-BSA 8-carbonyloctyl 4-O-(3,6-di-O-methyl--d-glucopyranosyl)--l-rhan pyranoside-BSA     

Metal resistance in Acinetobacter and its relation to β-lactamase production

Thirty nine clinical isolates of Acinetobacter belonging to six species were tested for resistance to 20 metal ions and their ability to produce -lactamase. Fifty two percent of the strains produced -lactamase. -Lactamase producers and non-producers were almost equally distributed in the different species. A. baumannii was the predominant biotype and was found to be most resistant to metals. Resistance to mercury was prevalent in -lactamase-producing A. baumannii only. Silver resistant strains of A. baumannii produced -lactamase. Sensitivity and resistance to copper and cadium was equally distributed between -lactamase producers and non-producers. -Lactamase-producer and -non-producer strains were uniformly sensitive to cadmium except Acinetobacter genospecies 1.     

Characterization of three endophytic, indole-3-acetic acid-producing yeasts occurring in Populus trees

Three endophytic yeast, one isolated from stems of wild cottonwood (Populus trichocarpa), two from stems of hybrid poplar (P. trichocarpa × Populus deltoides), were characterized by analyzing three ribosomal genes, the small subunit (18S), internal transcribed spacer (ITS), and D1/D2 region of the large subunit (26S). Phenotypic characteristics of the yeast isolates were also obtained using a commercial yeast identification kit and used for assisting the species identification. The isolate from wild cottonwood was identified to be closest to species Rhodotorula graminis. The two isolates from hybrid poplar were identified to be species Rhodotorula mucilaginosa. In addition, the three yeast isolates were observed to be able to produce indole-3-acetic acid (IAA), a phytohormone which can promote plant growth, when incubated with l-tryptophan. To our knowledge, the yeast strains presented in this study were the first endophytic yeast strains isolated from species of Populus.     

Production of β-fructofuranosidase byAspergillus japonicus

A newly isolated strain, MU-2, which produces very high -fructofuranosidase activity, was identified asAspergillus japonicus. For enzyme production by the strain, sucrose at 20% (w/v) was the best carbon source and yeast extract at 1.5 to 3% (w/v) the best nitrogen source. Total enzymatic activity and cell growth were at maximum after 48 h, at 1.57×10⁴ U/flask and 0.81 g dry cells/flask, respectively. The optimum pH value of the enzymatic reaction was between 5.0 and 5.5 and the optimum temperature 60 to 65°C. The enzyme produced 1-kestose (O--d-fructofuranosyl-(21)--d-fructofuranosyl -d-glucopyranoside) and nystose (O--d-fructofuranosyl-(21)--d-fructofuranosyl-(21)--d-fructofuranosyl -d-glucopyranoside) from sucrose by fructosyl-transferring activity. The strain was found to be very useful for industrial production of -fructofuranosidase.     

The molecular genetic differentiation of cultured <Emphasis Type="Italic">Saccharomyces</Emphasis> strains

A comparative molecular genetic study of cultured Saccharomyces strains isolated from the surface of berries and various fermentation processes showed that bakers yeast and black-currant isolates contain not only Saccharomyces cerevisiae but also S. cerevisiae × S. bayanus var. uvarum hybrids. The molecular karyotyping of bakers, brewers, and wine yeasts showed their polyploidy. The restriction enzyme analysis of noncoding rDNA regions (5.8S-ITS and IGS2) makes it possible to differentiate species of the genus Saccharomyces and to identify interspecies hybrids. The microsatellite primer (GTG)₅ can be used to study the populations of cultured S. cerevisiae strains.__________Translated from Mikrobiologiya, Vol. 74, No. 2, 2005, pp. 215–223.Original Russian Text Copyright © 2005 by Naumova, Zholudeva, Martynenko, Naumov.     

Identification of aryl-phospho-β-<Emphasis Type="SmallCaps">d</Emphasis>-glucosidases in<Emphasis Type="Italic"> Bacillus subtilis</Emphasis>

Four aryl-phospho--d-glucosidases were identified in Bacillus subtilis by using 4-methylumbelliferyl-phospho--d-glucopyranoside as a substrate. Two of these enzymes are the products of the bglA and bglH genes, previously suggested to encode aryl-phospho--d-glucosidases, while the other enzymes are encoded by the yckE and ydhP genes. Together, these four genes account for >99.9% of the glucosidase activity in B. subtilis on aryl-phospho--d-glucosides. yckE was expressed at a low and constant level during growth, sporulation, and spore germination, and was not induced by aryl--d-glucosides. ydhP was also not induced by aryl--d-glucosides. However, while ydhP was expressed at only a very low level in exponential-phase cells and germinating spores, this gene was expressed at a higher levels upon entry into the stationary phase of growth. Strains lacking yckE or ydhP exhibited no defects in growth, sporulation, or spore germination or in growth on aryl--d-glucosides. However, a strain lacking bglA, bglH and yckE grew poorly if at all on aryl--d-glucosides as the sole carbon source.Abbreviations MU 4-Methylumbelliferone - MUG 4-Methylumbelliferyl--d-glucopyranoside - MUGal 4-Methylumbelliferyl--d-galactopyranoside - MUG-P 4-Methylumbelliferyl--d-glucopyranoside-6-phosphate     

Interaction of the salivary low-molecular-weight mucin (MG2) with Actinobacillus actinomycetemcomitans

Periodontitis is associated with the presence of certain Gram-negative bacteria in the oral cavity, among these Actinobacillus actinomycetemcomitans. In order to determine which types of salivary components interact with A. actinomycetemcomitans two strains (HG 1175 and FDC Y4) were incubated with whole saliva and individual glandular secretions, viz. parotid, submandibular, and sublingual saliva. Immunochemical analysis by immunoblotting of bacteria-bound salivary proteins showed that IgA, the low-molecular mucin MG2, parotid agglutinin, and a 300 kDa sublingual and submandibular glycoprotein, were bound to the bacterial strains tested. In addition, adherence of A. actinomycetemcomitans to salivary proteins in a solid-phase was studied. After electrophoresis and transfer of salivary proteins to nitrocellulose membranes A. actinomycetemcomitans adhered only to MG2. In this assay periodate treatment, mild acid hydrolysis or neuraminidase digestion of the saliva glycoproteins abolished binding of two clinical isolates (HG 1175 and NY 664), suggesting that sialic acid residues on MG2 are involved in the binding. In contrast, adherence of the smooth laboratory strain Y4 was not affected by removal of sialic acid residues or even periodate treatment of MG2.Abbreviations S-IgA Secretory IgA - MG1 high-molecular-weight mucin - MG2 low-molecular-weight mucin - EP-GP extra parotid-glycoprotein - PRPs proline-rich proteins - SNA Sambucus nigra agglutinin - MAA Maackia amurensis agglutinin - PNA peanut agglutinin - UEA Ulex europaeus agglutinin     

Revision der GattungLoxonia (Gesneriaceae)

Within the genusLoxonia Jack, currently regarded as monotypic, three species are recognized:L. hirsuta Jack (Sumatra, Mentawai-Islands, Java, Borneo, Anambas-Islands, Malay Peninsula),L. discolor Jack (Sumatra) andL. burttiana A. Weber, spec. nova (Borneo). [Key with English translation p. 203.] There is evidence thatL. discolor is the most primitive species within the genus, the two others being derived from it.

Teil V der Beiträge zur Morphologie und Systematik derKlugieae undLoxonieae (Gesneriaceae).