Detection of Mycobacterium avium subsp. paratuberculosis in non-human primates.

BACKGROUND: Due to a sporadic occurrence of Mycobacterium avium subsp. paratuberculosis (MAP) in non-human primates (NHP), the susceptibility of different NHP to MAP should be investigated. METHODS: Fecal and tissue samples (ileum, ileocecal lymph node, bone marrow) of 20 animals (seven species) were analyzed by IS900-based PCRs and sequenced. Samples of MAP PCR positive NHP were further cultivated. RESULTS: MAP DNA was detectable in two animals; the ileum of a cottontop tamarin and the bone marrow of a common marmoset. Cultivation of MAP failed. Sequence analysis revealed 100% homology to the MAP-K10 sequence. Pathohistological examinations offered no direct correlation to a MAP infection. CONCLUSIONS: MAP was detected for the first time in a common marmoset. But as both NHP suffered from other diseases, an asymptomatic infection with MAP was assumed. The detection of MAP in the bone marrow might play a role in establishing latent paratuberculosis, as known from tuberculosis.

Rab11-FIP1A regulates early trafficking into the recycling endosomes.

The Rab11 family of small GTPases, along with the Rab11-family interacting proteins (Rab11-FIPs), are critical regulators of intracellular vesicle trafficking and recycling. We have identified a point mutation of Threonine-197 site to an Alanine in Rab11-FIP1A, which causes a dramatic dominant negative phenotype when expressed in HeLa cells. The normally perinuclear distribution of GFP-Rab11-FIP1A was condensed into a membranous cisternum with almost no GFP-Rab11-FIP1A(T197A) remaining outside of this central locus. Also, this condensed GFP-FIP1A(T197A) altered the distribution of proteins in the Rab11a recycling pathway including endogenous Rab11a, Rab11-FIP1C, and transferrin receptor (CD71). Furthermore, this condensed GFP-FIP1A(T197A)-containing structure exhibited little movement in live HeLa cells. Expression of GFP-FIP1A(T197A) caused a strong blockade of transferrin recycling. Treatment of cells expressing GFP-FIP1A(T197A) with nocodazole did not disperse the Rab11a-containing recycling system. We also found that Rab5 and EEA1 were accumulated in membranes by GFP-Rab11-FIP1A but Rab4 was unaffected, suggesting that a direct pathway may exist from early endosomes into the Rab11a-containing recycling system. Our study of a potent inhibitory trafficking mutation in Rab11-FIP1A shows that Rab11-FIP1A associates with and regulates trafficking at an early step in the process of membrane recycling.

DETECTION OF MYCOBACTERIUM AVIUM SUBSPECIES PARATUBERCULOSIS IN ROCK HYRAXES ( PROCAVIA CAPENSIS) IMPORTED FROM SOUTH AFRICA.

Mycobacterium avium subspecies paratuberculosis (MAP) causes chronic, progressive, and consecutively fatal enteritis, especially in ruminants. MAP distribution among wildlife is not yet clear. In this study, three wild-born rock hyraxes ( Procavia capensis) had been imported from South Africa to a German zoological garden. During the quarantine period, four young animals were born. The wild-born animals showed symptoms of mild diarrhea shortly after their arrival in the zoological garden, but all routine parasitological and bacteriologic tests performed were negative. Therefore, the animals were additionally tested for MAP infection. MAP DNA was detected by seminested PCR (snPCR) in a pooled fecal sample of the seven animals. Subsequent PCR analysis of the individual feces samples confirmed the excretion of MAP in two rock hyraxes (one wild-born and one born in captivity). Sequence analysis of the corresponding 278-bp amplicons revealed 100% homology to the reference MAP-K10 IS900 sequence. No antibody response against MAP was detected in the individual serum samples. MAP-specific postmortem lesions were not observed by gross pathology and histology, neither after death nor after euthanization of the animals. Nevertheless, MAP was detected by snPCR and culture in the gastrointestinal tract, urogenital tract, cardiovascular system, and/or respiratory system of three other animals of the group (one wild-born and two born in captivity). This study is the first report confirming MAP occurrence in rock hyraxes. Therefore, it is recommended that veterinarians and zoo employees consider rock hyraxes as a possible source of MAP infection for domestic livestock in South Africa and the valuable animal stock of zoological facilities.

Rab11-FIP2 interaction with MYO5B regulates movement of Rab11a-containing recycling vesicles.

A tripartite association of Rab11a with both Rab11-FIP2 and MYO5B regulates recycling endosome trafficking. We sought to define the intermolecular interactions required between Rab11-FIP2 and MYO5B. Using a random mutagenesis strategy, we identified point mutations at S229P or G233E in Rab11-FIP2 that caused loss of interaction with MYO5B in yeast two-hybrid assays as well as loss of interaction of Rab11-FIP2(129-356) with MYO5B tail when expressed in HeLa cells. Single mutations or the double S229P/G233E mutation failed to alter the association of full-length Rab11-FIP2 with MYO5B tail in HeLa cells. While EGFP-Rab11-FIP2 wild type colocalized with endogenous MYO5B staining in MDCK cells, EGFP-Rab11-FIP2(S229P/G233E) showed a significant decrease in localization with endogenous MYO5B. Analysis of Rab11a-containing vesicle movement in live HeLa cells demonstrated that when the MYO5B/Rab11-FIP2 association is perturbed by mutation or by Rab11-FIP2 knockdown, vesicle movement is increased in both speed and track length, consistent with an impairment of MYO5B tethering at the cytoskeleton. These results support a critical role for the interaction of MYO5B with Rab11-FIP2 in stabilizing the functional complex with Rab11a, which regulates dynamic movements of membrane recycling vesicles.

Detection of airborne bacteria in a duck production facility with two different personal air sampling devices for an exposure assessment.

Prevalent airborne microorganisms are not well characterized in industrial animal production buildings with respect to their quantity or quality. To investigate the work-related microbial exposure, personal bioaerosol sampling during the whole working day is recommended. Therefore, bioaerosol sampling in a duck hatchery and a duck house with two personal air sampling devices, a filter-based PGP and a NIOSH particle size separator, was performed. Subsequent, quantitative and qualitative analyses were carried out with" culture independent methods. Total cell concentrations (TCC) determined via fluorescence microscopy showed no difference between the two devices. In average, 8 × 10(6) cells/m(3) were determined in the air of the duck hatchery and 5 × 10(7) cells/m(3) in the air of the duck house. A Generated Restriction Fragment Length Polymorphism (RFLP) pattern revealed deviant bacterial compositions comparing samples collected with both devices. Clone library analyses based on 16S rRNA gene sequence analysis from the hatchery's air showed 65% similarity between the two sampling devices. Detailed 16S rRNA gene sequence analyses showed the occurrence of bacterial species like Acinetobacter baumannii, Enterococcus faecalis, Escherichia sp., and Shigella sp.; and a group of Staphylococcus delphini, S. intermedius, and S. pseudintermedius that provided the evidence of potential exposure to risk group 2 bacteria at the hatchery workplace. Size fractionated sampling with the developed by the Institute for Occupational Safety and Health of the German Social Accident Insurance (IFA) device revealed that pathogenic bacteria would deposit in the inhalable, the thorax, and possibly alveolar dust fraction according to EN481. TCC analysis showed the deposition of bacterial cells in the third stage (< 1µm) at the NIOSH device which implies that bacteria can reach deep into the lungs and contaminate the alveolus after inhalation. Nevertheless, both personal sampling devices could be recommended for exposure assessment at agricultural workplaces.

Core Exchange: A Professional Development Program for Shared Resource Personnel.

Introduction/Objective: Academic institutions often struggle to meet the unique professional development needs of shared resource personnel, who require business skills, project and people management expertise, and an active, collaborative network of shared resource colleagues. Materials and Methods: We launched the Vanderbilt Core Exchange professional development and networking program in 2020. The program was intentionally designed with core personnel input and supports faculty and staff from more than 80 shared resources across Vanderbilt University and Vanderbilt University Medical Center. Resources offered include a quarterly seminar series with both virtual and in-person events, a website for accessing professional development materials and session recordings, and a dedicated online discussion group for core personnel networking. Results: There have been 11 Vanderbilt Core Exchange events to date: 2 in person and 9 virtual. In-person events averaged 35 attendees, and virtual events averaged 45 attendees. Topics included equipment grant writing, marketing, handling difficult conversations, managing different workplace work styles, communication and project management tools, and the importance of self-care. Survey responses collected after each event were highly positive and informed areas of improvement and future event topics. Discussion: This model of local shared resource professional development serves as a template for institutions who desire to create opportunities for collaboration and community building. With a small coordinating committee of dedicated individuals, an institution-wide professional development and networking program can be successfully established even with limited resources.

Quantification of Saccharopolyspora rectivirgula in composting plants: assessment of the relevance of S. rectivirgula.

Exposure to bioaerosols in composting plants can lead to negative health effects on compost workers. Health complaints vary between cough, irritation of the eyes and the skin, sinusitis, or dyspnea among others. It is fact that compost materials harbor high concentrations of microorganisms, which were aerosolized during handling compost. Within the present study, total cell numbers between 3.4 × 10(4) and 1.6 × 10(8) cell counts per m(3) air were determined after 4',6-Diamidin-2-phenylindol DAPI staining in 124 samples from German composting plants. Special attention should be paid to some specific microorganisms, which are able to cause health complaints. Saccharopolyspora rectivirgula, known to be one of the major causes of extrinsic allergic alveolitis (EAA, also called hypersensitivity pneumonitis, HP), was often found in environments of agricultural production, where the classical form of EAA ('farmer's lung disease') is common, but also in composting plants. In Germany, cases are known where workers had to terminate their work due to this disease. However, up to now, the relevance of S. rectivirgula at composting plants is unexplained. This study showed that high concentrations of airborne S. rectivirgula were found in composting plants similar to that found in agricultural production. Altogether, in 86.7% of the 124 analyzed samples, S. rectivirgula was detected using quantitative real-time polymerase chain reaction (PCR). Estimated concentrations ranged between 1.24 × 10(2) cell counts of S. rectivirgula per cubic meter air next to the rotted residues and 1.5 × 10(7) cell counts next to a converter. Furthermore, our methodical proceedings were verified. To analyze DNA extraction limits through the amount of cells within one sample, the DNA concentration was compared with total cell counts (TCCs). Altogether, when TCC was <1.4 × 10(5) cells per DNA extraction assay, no DNA was measurable; when TCC reached 3.5 × 10(6) cells, DNA was always detectable by fluorometric method. To overcome limitation of DNA measurement using fluorometric method, samples without measurable DNA were inserted in a PCR assay with universal primers. Results showed that a gain of 37% was possible, when samples were additionally analyzed by universal PCR. Hence, cell counts >2.0 × 10(6) cells were necessary to measure DNA concentrations in 90% of the analyzed samples, whereas cell counts <3.0 × 10(5) are sufficient to detect PCR products. Therefore, sampling of bioaerosols should be done in consideration of the expected cell count per cubic meter air. Note, to get measurable DNA using a fluorometer, >3.5 × 10(6) cells must be sampled for one DNA extraction assay. With this study, the real-time PCR approach for the detection of S. rectivirgula at workplaces in compost plants was revised, and the results revealed that this method is suitable for occupational exposure measurements.

Connect, Collaborate, and Succeed: The Ninth Annual Meeting of the Southeastern Association of Shared Resources (SEASR) Nashville, TN, USA, June 8-10, 2022.

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Neutrophil trafficking to the site of infection requires Cpt1a-dependent fatty acid ß-oxidation.

Cellular metabolism influences immune cell function, with mitochondrial fatty acid ß-oxidation and oxidative phosphorylation required for multiple immune cell phenotypes. Carnitine palmitoyltransferase 1a (Cpt1a) is considered the rate-limiting enzyme for mitochondrial metabolism of long-chain fatty acids, and Cpt1a deficiency is associated with infant mortality and infection risk. This study was undertaken to test the hypothesis that impairment in Cpt1a-dependent fatty acid oxidation results in increased susceptibility to infection. Screening the Cpt1a gene for common variants predicted to affect protein function revealed allele rs2229738_T, which was associated with pneumonia risk in a targeted human phenome association study. Pharmacologic inhibition of Cpt1a increases mortality and impairs control of the infection in a murine model of bacterial pneumonia. Susceptibility to pneumonia is associated with blunted neutrophilic responses in mice and humans that result from impaired neutrophil trafficking to the site of infection. Chemotaxis responsible for neutrophil trafficking requires Cpt1a-dependent mitochondrial fatty acid oxidation for amplification of chemoattractant signals. These findings identify Cpt1a as a potential host determinant of infection susceptibility and demonstrate a requirement for mitochondrial fatty acid oxidation in neutrophil biology.

Detection of Saccharopolyspora rectivirgula by quantitative real-time PCR.

The thermophilic actinomycete species Saccharopolyspora rectivirgula has been associated with the exogen allergic alveolitis (EAA). EAA is caused by the inhalation of high amounts of airborne spores that can be found for example in environments of agricultural production, compost facilities, mushroom cultivation rooms, or rooms with technical air moistening. Because of the medical relevance of S. rectivirgula, a reliable detection system is needed. Therefore, a quantitative real-time polymerase chain reaction (qPCR) primer system was designed, targeting the 16S rRNA gene of the type strain S. rectivirgula DSM 43747(T) and six other S. rectivirgula reference strains. Our investigation showed that S. rectivirgula presumably own four operons of the 16S rRNA gene, which has to be considered for estimation of cell equivalents. Furthermore, the DNA recovery efficiency from these strains was tested in combination with bioaerosol or material sample as well as the influence of non-target DNA to the recovery rate. Results showed a recovery DNA efficiency of 7-55%. The recovery rate of DNA in a mixture with non-target DNA resulted in ∼87%. In summary, a high amplification efficiency using real-time PCR was found, for which estimated concentrations revealed cell numbers of 2.7 × 10(5) cells m(-3) in bioaerosol and 2.8 × 10(6) cells g(-1) fw(-1) in material samples from a duck house. The specificity of the new developed quantification system was shown by generation of two clone libraries from bioarosol samples, from a duck house, and from a composting plant. Totally, the results clearly show the specificity and practicability of the established qPCR assay for detection of S. rectivirgula.

D-AKAP2 interacts with Rab4 and Rab11 through its RGS domains and regulates transferrin receptor recycling.

Dual-specific A-kinase-anchoring protein 2 (D-AKAP2/AKAP10), which interacts at its carboxyl terminus with protein kinase A and PDZ domain proteins, contains two tandem regulator of G-protein signaling (RGS) domains for which the binding partners have remained unknown. We show here that these RGS domains interact with Rab11 and GTP-bound Rab4, the first demonstration of RGS domains binding small GTPases. Rab4 and Rab11 help regulate membrane trafficking through the endocytic recycling pathways by recruiting effector proteins to specific membrane domains. Although D-AKAP2 is primarily cytosolic in HeLa cells, a fraction of the protein localizes to endosomes and can be recruited there to a greater extent by overexpression of Rab4 or Rab11. D-AKAP2 also regulates the morphology of the Rab11-containing compartment, with co-expression causing accumulation of both proteins on enlarged endosomes. Knockdown of D-AKAP2 by RNA interference caused a redistribution of both Rab11 and the constitutively recycling transferrin receptor to the periphery of cells. Knockdown also caused an increase in the rate of transferrin recycling, suggesting that D-AKAP2 promotes accumulation of recycling proteins in the Rab4/Rab11-positive endocytic recycling compartment.

Sensory ciliogenesis in Caenorhabditis elegans: assignment of IFT components into distinct modules based on transport and phenotypic profiles.

Sensory cilium biogenesis within Caenorhabditis elegans neurons depends on the kinesin-2-dependent intraflagellar transport (IFT) of ciliary precursors associated with IFT particles to the axoneme tip. Here we analyzed the molecular organization of the IFT machinery by comparing the in vivo transport and phenotypic profiles of multiple proteins involved in IFT and ciliogenesis. Based on their motility in wild-type and bbs (Bardet-Biedl syndrome) mutants, IFT proteins were classified into groups with similar transport profiles that we refer to as "modules." We also analyzed the distribution and transport of fluorescent IFT particles in multiple known ciliary mutants and 49 new ciliary mutants. Most of the latter mutants were snip-SNP mapped and one, namely dyf-14(ks69), was cloned and found to encode a conserved protein essential for ciliogenesis. The products of these ciliogenesis genes could also be assigned to the aforementioned set of modules or to specific aspects of ciliogenesis, based on IFT particle dynamics and ciliary mutant phenotypes. Although binding assays would be required to confirm direct physical interactions, the results are consistent with the hypothesis that the C. elegans IFT machinery has a modular design, consisting of modules IFT-subcomplex A, IFT-subcomplex B, and a BBS protein complex, in addition to motor and cargo modules, with each module contributing to distinct functional aspects of IFT or ciliogenesis.

Thermophilic methanogenic Archaea in compost material: occurrence, persistence and possible mechanisms for their distribution to other environments.

Since compost is widely used as soil amendment and the fact that during the processing of compost material high amounts of microorganisms are released into the air, we investigated whether compost may act as a carrier for thermophilic methanogens to temperate soils. All eight investigated compost materials showed a clear methane production potential between 0.01 and 0.98 micromol CH(4) g dw(-1)h(-1) at 50 degrees C. Single strand conformation polymorphism (SSCP) and cloning analysis indicated the presence of Methanosarcina thermophila, Methanoculleus thermophilus, and Methanobacterium formicicum. Bioaerosols collected during the turning of a compost pile showed both a highly similar SSCP profile compared to the corresponding compost material and clear methane production during anoxic incubation in selective medium at 50 degrees C. Both observations indicated a considerable release of thermophilic methanogens into the air. To analyse the persistence of compost-borne thermophilic methanogens in temperate oxic soils, we therefore studied their potential activity in compost and compost/soil mixtures, which was brought to a meadow soil, as well as in an agricultural soil fertilised with compost. After 24h anoxic incubation at 50 degrees C, all samples containing compost showed a clear methanogenic activity, even 1 year after application. In combination with the in vitro observed resilience of the compost-borne methanogens against desiccation and UV radiation we assume that compost material acts as an effective carrier for the distribution of thermophilic methanogens by fertilisation and wind.

XBX-1 encodes a dynein light intermediate chain required for retrograde intraflagellar transport and cilia assembly in Caenorhabditis elegans.

Intraflagellar transport (IFT) is a process required for flagella and cilia assembly that describes the dynein and kinesin mediated movement of particles along axonemes that consists of an A and a B complex, defects in which disrupt retrograde and anterograde transport, respectively. Herein, we describe a novel Caenorhabditis elegans gene, xbx-1, that is required for retrograde IFT and shares homology with a mammalian dynein light intermediate chain (D2LIC). xbx-1 expression in ciliated sensory neurons is regulated by the transcription factor DAF-19, as demonstrated previously for genes encoding IFT complex B proteins. XBX-1 localizes to the base of the cilia and undergoes anterograde and retrograde movement along the axoneme. Disruption of xbx-1 results in cilia defects and causes behavioral abnormalities observed in other cilia mutants. Analysis of cilia in xbx-1 mutants reveals that they are shortened and have a bulb like structure in which IFT proteins accumulate. The role of XBX-1 in IFT was further confirmed by analyzing the effect that other IFT mutations have on XBX-1 localization and movement. In contrast to other IFT proteins, retrograde XBX-1 movement was detected in complex A mutants. Our results suggest that the DLIC protein XBX-1 functions together with the CHE-3 dynein in retrograde IFT, downstream of the complex A proteins.

Heterogeneity in Cultivation-Based Monitoring of Airborne Bacterial Biodiversity in Animal Farms.

Diversity analyses of bioaerosol samples from highly loaded workplaces as found in agricultural production or waste management help to improve the knowledge of exposure levels of workers. However, different used methods resulting in the detection of different bacterial species at the same work places. The present study obviously supports the deviation of received results using cultivation and further isolation approaches. Within the present study, the bacterial community at workplaces was estimated using the powerful tool of 16S rRNA gene sequence analyses after cultivation procedure. To avoid complex isolation procedures, the suitability of cultivation and subsequent cloning procedures was determined in bioaerosols from a duck hatchery. Diversity analysis of one bioaerosol sample, which was prepared independently three times in parallel, resulted in similarity values of 38.5%-57.1%. Further, similarity analysis calculated from three independent bioaerosol samplings on 1 day resulted in 31%-40% similarity. Although similar concentration between 2.22 × 106 and 4.46 × 106 CFU per m3 hatchery air were measured, in a ring-like trail, diversity analyses from six labs differ widely, resulting in 38.9%-78.6% divergence. The present method seems to be very useful for diversity analysis of bioaerosol samples, although heterogeneity in monitoring of airborne bacteria via cultivation was pointed out.

Preliminary Validation of a Method Combining Cultivation and Cloning-Based Approaches to Monitor Airborne Bacteria.

It is already known that occupational exposure to bioaerosols or organic dust could be harmful for occupants, but mostly the correlation to occurring bacteria is missing. Especially, cultivation of bacteria from bioaerosols is important to get an insight on occurring and possibly infectious bacteria. These measures are highly time consuming and cost intensive. Therefore, to monitor bacterial diversity in bioaerosol samples and to avoid isolation procedures, an approach was applied using a combination of cultivation and cloning-based approach. Preliminary validation of the method was determined using 11 different bacterial strains. After DNA extraction and 16S rRNA gene amplification of grown colonies, subsequent cloning and sequencing was conducted. Initially, to figure out a suitable DNA extraction method, applicable for different airborne bacteria, four DNA extraction protocols were compared. Significantly, best results were determined using the FAST DNA®Spin Kit for Soil with respect to DNA quantity and quality of bacterial cultures. Cloning approach from a mixture of amplified 16S rRNA genes of 11 isolates and following sequence data analysis shows a recovery of all strains when five clones per bacterial strain were analysed. The results clearly demonstrate that a combination of cultivation-based approaches and cloning processes can simplify bioaerosol monitoring of viable and probably infectious bacteria. The implementation of the present method into practice allows a simple and preventive investigation of bioaerosols at work places.

Development of a Recombinase Polymerase Amplification Assay for Rapid Detection of the Mycobacterium avium subsp. paratuberculosis.

BACKGROUND: The detection of Mycobacterium avium subsp. paratuberculosis (MAP) infections in ruminants is crucial to control spread among animals and to humans. Cultivation of MAP is seen as the gold standard for detection, although it is very time consuming and labour intensive. In addition, several PCR assays have been developed to detect MAP in around 90 minutes, but these assays required highly sophisticated equipment as well as lengthy and complicated procedure. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we have developed a rapid assay for the detection of MAP based on the recombinase polymerase amplification (RPA) assay targeting a MAP specific region, the IS900 gene. The detection limit was 16 DNA molecules in 15 minutes as determined by the probit analysis on eight runs of the plasmid standard. Cross reactivity with other mycobacterial and environmentally associated bacterial strains was not observed. The clinical performance of the MAP RPA assay was tested using 48 MAP-positive and 20 MAP-negative blood, sperm, faecal and tissue samples. All results were compared with reads of a highly sensitive real-time PCR assay. The specificity of the MAP RPA assay was 100%, while the sensitivity was 89.5%. CONCLUSIONS/SIGNIFICANCE: The RPA assay is quicker and much easier to handle than real-time PCR. All RPA reagents were cold-chain independent. Moreover, combining RPA assay with a simple extraction protocol will maximize its use at point of need for rapid detection of MAP.

Prauserella muralis sp. nov., from the indoor environment.

A novel Gram-stain-positive, mycelium-forming actinobacterium (05-Be-005(T)) isolated from the wall of an indoor environment was studied for its taxonomic position. The isolated strain formed a substrate mycelium that fragmented into rod-shaped cells and showed an aerial mycelium on medium M79. On the basis of 16S rRNA gene sequence similarity studies, strain 05-Be-005(T) was shown to belong to the genus Prauserella, closely related to Prauserella rugosa DSM 43194(T) (96.6 % similarity), Prauserella alba YIM 90005(T) (95.9 %) and Prauserella halophila YIM 90001(T) (95.4 %). The predominant menaquinone was MK-9(H(4)); whole-cell hydrolysates contained meso-diaminopimelic acid as the diagnostic diamino acid of the cell wall and arabinose and galactose as the main sugars. Mycolic acids were absent. The polar lipid profile consisted of the lipids diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylmethylethanolamine, phosphatidylinositol, phosphatidylserine and an unknown phospholipid. Major fatty acids C(16 : 0) iso, C(16 : 0), C(17 : 1)omega8c and C(17 : 1)omega6c supported the affiliation of strain 05-Be-005(T) to the genus Prauserella. The results of physiological and biochemical tests allowed clear phenotypic differentiation of strain 05-Be-005(T) from the three known Prauserella species. Strain 05-Be-005(T) represents a novel Prauserella species, for which we propose the name Prauserella muralis sp. nov., with the type strain 05-Be-005(T) (=CCUG 57426(T) =NRRL B-24780(T) =CCM 7635(T)=DSM 45305(T)).

Citricoccus parietis sp. nov., isolated from a mould-colonized wall and emended description of Citricoccus alkalitolerans Li et al. 2005.

A Gram-positive, coccoid-shaped organism (strain 02-Je-010(T)), forming yellow-pigmented colonies was isolated from the wall of an indoor environment. On the basis of 16S rRNA gene sequence similarity studies, it was shown that strain 02-Je-010(T) belongs to the genus Citricoccus with sequence similarities of 98.9 % to Citricoccus alkalitolerans DSM 15665(T) and 98.6 % to Citricoccus muralis DSM 14442(T). Cell-wall sugars were mannose and glucose. The diagnostic diamino acid of the peptidoglycan was lysine. The major menaquinones detected were MK-9(H(2)) and MK-8(H(2)). The polar lipid profile consisted of the major lipids diphosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol and moderate amounts of two unknown phospholipids and two unknown glycolipids. The fatty acid profile comprised major amounts of anteiso-C(15 : 0), anteiso-C(17 : 0) and iso-C(15 : 0). All these data supported the affiliation of strain 02-Je-010(T) to the genus Citricoccus. The results of DNA-DNA hybridization and physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain 02-Je-010(T) from the two recognized Citricoccus species. For these reasons, strain 02-Je-010(T) represents a novel species, for which the name Citricoccus parietis sp. nov. is proposed, with the type strain 02-Je-010(T) (=CCUG 57388(T)=CCM 7609(T)).