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Home , Nitrosomonadales, Nitrosomonas eutropha, Paracoccus, Paracoccus denitrificans, Pseudomonas monteilii, Rhodobacteraceae

APPLICATION FORM DETERMINATION

Determine if an organism is a new organism under the Hazardous Substances and New Organisms Act 1996

Send by post to: Environmental Protection Authority, Private Bag 63002, Wellington 6140 OR email to: [email protected]

Application number

APP201895

Applicant

Neil Pritchard

Key contact

NPN Ltd

www.epa.govt.nz 2

Application to determine if an organism is a new organism

Important

This application form is used to determine if an organism is a new organism. If you need help to complete this form, please look at our website (www.epa.govt.nz) or email us at [email protected]. This application form will be made publicly available so any confidential information must be collated in a separate labelled appendix. The fee for this application can be found on our website at www.epa.govt.nz. This form was approved on 1 May 2012.

May 2012 EPA0159 3

Application to determine if an organism is a new organism

1. Information about the new organism

What is the name of the new organism? Briefly describe the biology of the organism. Is it a genetically modified organism?

Pseudomonas monteilii

Kingdom:

Phylum:

Class: Gamma Proteobacteria

Order:

Family:

Genus:

Species: Pseudomonas monteilii Elomari et al., 1997

Binomial name: Pseudomonas monteilii Elomari et al., 1997. Pseudomonas monteilii is a Gram-negative, rod- shaped, motile bacterium isolated from human bronchial aspirate (Elomari et al 1997). They are incapable of liquefing gelatin. They grow at 10°C but not at 41°C, produce fluorescent pigments, catalase, and cytochrome oxidase, and possesse the arginine dihydrolase system. They are capable of respiratory but not fermentative metabolism. (Masua et al 2007)

Rhodococcus pyridinivorans

Kingdom: Bacteria

Phylum: Actinobacteria

Class: Actinobacteria

Order: Actinomycetales

Family: Nocardiaceae

Genus: Rhodococcus

Species: Rhodococcus pyridinivorans Yoon et al., 2000

The cells are non-spore-forming, non-motile and Gram-positive, but are Gram-variable in old cultures. Substrate mycelia that are on the surface and penetrate into the agar media are visible and fragment into short rod-to- elements. The cells are rods and branched filaments during the early growth phase and then fragment into short rods or cocci. Colonies are light orange in colour, opaque and raised with slightly irregular edges on TSA. Colonies have irregularly round wrinkles. Grows well over a broad pH range (6.0-9.0); the optimal pH range is 7.5-8.5. Grows optimally at 30-37 °C; growth occurs at 10 and 45 °C but does not occur at 50 °C. Catalase- and urease- positive. Oxidase- and DNase-negative. Arbutin, aesculin, Tween 80, tyrosine and urea are hydrolysed. Casein,

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Application to determine if an organism is a new organism hypoxanthine, starch and xanthine are not hydrolysed. Nitrate is reduced to nitrite. Voges- Proskauer and methyl red reactions are negative. H2S is produced. Acid is produced from d-fructose, glycerol, d-mannitol, d-mannose, d- ribose, salicin, d-sorbitol and starch. No acid is produced from larabinose, D-cellobiose, D-galactose, D-glucose, inulin, lactose, maltose, D-raffinose, L-rhamnose, trehalose or D-xylose. Acetate, citrate, fumarate and succinate are utilized. Benzoate, formate, hippurate and tartrate are not utilized. Degrades pyridine. The cell wall contains meso-diaminopimelic acid, arabinose and galactose (wall chemotype IV). The predominant menaquinone is MK- 8(H2). The major fatty acids are C16:0, C18:1 cis9, 10-methyl-C18:1 (TBSA). Mycolic acids with 36-46 carbon atoms are present. The GC content of the DNA is 66 mol% (as determined by HPLC). (Yoon et al. 2000)

R. pyridinivorans has been characterized based on phylogenetic and chemotaxonomic descriptors such as G+C content, 16S rDNA sequences, and DNA-DNA relatedness. The 16S rDNA analysis revealed 99% nucleotide similarity to that of the type strain of R. rhodochrous, which is not known to be pathogenic.

Paracoccus pantotrophus

Kingdom: Bacteria

Phylum: Proteobacteria

Class:

Order:

Family:

Genus:

Species: Paracoccus pantotrophus (Robertson and Kuenen 1984) Rainey et al. 1999

Synonym: Thiosphaera pantotropha

Some strains are capable of aerobic (simultaneous reduction of and nitrate) and heterotrophic (oxidation of ammonium to nitrite during heterotrophic growth); some strains are capable of aerobic growth on formate, aerobic chemolithoautotrophic growth with carbon disulfide or as energy substrates, methylotrophic growth on or methylated sulfides, and heterotrophic growth on diethyl sulfide, thioethanol, thioacetic acid or substituted thiophenes; some strains can grow anaerobically with denitrification on thiosulfate, carbon disulfide, methanol or formate as energy sources; some strains contain plasmids of 85-110 kb in size and megaplasmids greater than 450 kb in size; distinction from other species of Paracoccus can be confirmed by comparison of 16S rRNA gene sequence and DNA hybridization; the amino acid sequence of its cytochrome c-550 differs by about 16% from that of P. denitrificans; and the GC content of the DNA is 64-68 mol% (type strain GC content is 66 mol%). Kelly et al (2006)

Nitrosomonas eutropha

Kingdom: Bacteria

Phylum: Proteobacteria

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Application to determine if an organism is a new organism

Class: Beta Proteobacteria

Order:

Family: Nitrosomonadaceae

Genus:

Species: Koops et al., 1991

Cells tend to be pleomorphic, rod to pear-shaped (sometimes coccoid). One or both ends pointed. Cells 1.0-1.3 by 1.6-2.3 μm in size, occasionally in short chains. Cells are motile. Carboxysomes present. UtiIization of urea not observed. No salt requirement, high tolerance of increasing concentrations. The G+C content of the DNA is 47.9-48.5 mol % (Tm).

The organism that is the subject of this application is also the subject of: a. an innovative medicine application as defined in section 23A of the Medicines Act 1981. Yes No b. an innovative agricultural compound application as defined in Part 6 of the Agricultural Compounds and Veterinary Medicines Act 1997. Yes No

2. What evidence do you have that the organism was present in New Zealand immediately prior to 29 July 1998?

Pseudomonas monteilii

The genus Pseudomonas encompasses the most diverse and ecologically significant group of bacteria on earth. Members of the genus are found in all of the major natural environments (terrestrial, freshwater and marine) and also form associations with plants and animals. This universal distribution suggests a remarkable degree of physiological and genetic adaptability (Spiers et al. 2000). Pseudomonas is a genus with widespread occurrence in water and in plant seeds such as dicots. Many members of the genus have been reported in New Zealand, including and Pseudomonas oryzihabitans (http://nzfungi2.landcareresearch.co.nz), which are taxonomically closely related with Pseudomonas monteilii.

Pseudomonas monteilii is a versatile bacterium found in various niches (Remold et al. 2011). Pseudomonas monteilii strains are associated with degradation of aromatic and heterocyclic compounds. P. monteiliiis is an aspirate found in many international hospitals, and where organisms are carried by people, it is easy to argue that they have been carried in to NZ even if they haven’t been recorded. Some Pseudomonas monteilii strains were isolated from clinical specimens, yet no P. monteilii infection has been reported. The P. monteilii strains are phenotypically and genotypically homogeneous and can be differentiated from related nonphytopathogenic fluorescent members of section I of the genus Pseudomonas.

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Application to determine if an organism is a new organism

The clinical significance of P. monteilii is not known. Future isolates of this new organism should be investigated to determine their role in nosocomial infections. At present, it is hypothesized that P. monteilii is a rare opportunistic or colonizer.

Pseudomonas spp. are ubiquitous in the environment. As opportunistic , some Pseudomonas spp. could invade the host tissue and cause infection and bacteremia in immunocompromised hosts. P. aeruginosa is the most common agent associated with infection and inflammation during contact lens wear. Other Pseudomonas species are also opportunistic; however, cases of infection are rare.

Rhodococcus pyridinivorans

Members of Rhodococcus have been found to thrive in a broad range of environments, including soil, water, and eukaryotic cells. Several species of the genus have been found in New Zealand, such as Rhodococcus equi, Rhodococcus fascians, and Rhodococcus rhodochrous (http://nzfungi2.landcareresearch.co.nz)

Members of the genus Rhodococcus are mostly soil saprophytes although they are common in many environments, and a large number of them are known as toxic-material decomposing strains. Rhodococcus pyridinivorans was originally isolated as an extremely efficient pyridine-degrading coryneform bacterium from industrial wastewater in Korea The species R. pyridinivorans comprises strains that are metabolically versatile and are able to degrade numerous aromatic compounds.

The pathogenicity of the bacteria that belonged to Rhodococcus was investigated based on information from the literature. Previous reports revealed that R. equi and R. facians are severely pathogenic for animals and plants, respectively. R. equi causes foal pneumonia and human opportunistic infections, while R. facians causes crown gall of tobacco leaves. In other studies, R. sputi caused boar tuberculosis and lymphadenitis, R. aurantiacus caused cerebral meningitis in a cancer patient, and R. erythropolis was detected in AIDS patients. However, since the identification of these three bacteria was incomplete, it is doubtful whether these bacteria are pathogenic

Some Rhodococcus species have been used in industrial processes. For example, R. rhodochrous produces citric acid and steroids, and Rhodococcus sp. converts benzonitrile to benzoic acid and produces glutamic acid. R. erythropolis is also used in acrylamide production and as a bacterial flocculant. These industrial applications suggest that most of the Rhodococcus species are non-pathogenic in plants and animals.

R. pyridinivorans is commonly present in contaminated soil or water with a variety of xenobiotics, specifically toxic aromatic compounds. These compounds are produced by both anthropogenic and natural activities and accumulate in the environment. Many these aromatic compounds, including pyridine, are toxic with implications for public and ecosystem health. These xenobiotic and organic compounds are more efficiently degraded by R. pyridinivorans bacteria than in the . Thus, their biodegrading capabilities of the toxic compounds contribute greatly to . This species has not been reported to being pathogenic.

Paracoccus pantotrophus

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Application to determine if an organism is a new organism

Paracoccus pantotrophus has been isolated in NZ from waste treatment plants. Members of Paracoccus are common soil bacteria. Tens of species belonging to the genus have been recorded in New Zealand

Paracoccus spp. are capable of reducing nitrate to nitrogen gas and are widely found in nitrogen rich environments. As reported by Foglar et al. (2005) Paracoccus was the dominant bacterial species found after wastewater from an industrial yeast process was biologically treated.

Except for two human infections by Paracoccus yeei reported in the literature (Funke et al. 2004), there have been no reports associating the genus Paracoccus with clinical infections in either humans or animals.

Paracoccus pantotrophus is not a known pathogen, and no other concerns have been identified either in the limited literature available.

Nitrosomonas eutropha

Habitat: common in municipal .and industrial, sewage disposal systems; seems to be distributed generally in strongly eutrophic environments

Nitrosomonas europaea and Nitrosomonas eutropha have been recorded in New Zealand (Silyn-Roberts and Lewis 2001)

The strains of Nitrosamines eutropha are commonly isolated from municipal and industrial sewage disposal systems; they seem to be distributed generally in strongly eutrophic environments (those rich in organic nutrients and minerals). Nitrosomonas eutropha also has a high tolerance for elevated ammonia concentrations. Nitrosomonas eutropha is able to grow anaerobically, utilizing nitrite as an electron acceptor and H2 as a reductant. As an ammonia-oxidizing bacterium, Nitrosomonas eutropha is believed to contribute significantly to the global production of , a product of nitrite reduction.

Nitrosomonas eutropha is not known to have any pathogenic qualities. This species is a common environmental bacterium found in New Zealand’s aquatic and terrestrial ecosystems. This species has also been commonly researched in both pure and mixed cultures

Given the wide distribution of microbes in the environment, there is no reason to think these four are not already present in New Zealand.

3. Appendices(s) and referenced material (if any) and glossary (if required)

Foglar, L., Briski, F., Sipos, L and Vukovic, M (2005). High nitrate removal from synthetic wastewater with the mixed bacterial culture. Bioresource Technology 96 (8): 879-888.

Funke, G., Frodl, R and Sommer, H. (2004). First comprehensively documented case of Paracoccus yeei in a human. Journal of Clinical Microbiology 42: 3366-3368

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Application to determine if an organism is a new organism

Kelly, D.P., Rainey, F.A and Wood, A, P. (2006). The genus Paracoccus. Prokaryotes 5: 232-249.

Masuda, M., Yamasaki Y., Uena, S and Inoue, A. (2007). Isolation of bisphenol A-tolerant/degrading Pseudomonas monteilii starin N-502. 11: 355-362

Rainey, F.A., Kelly, D.P., Stackebrandt, E, Burghardt, J, Hiraishi, A., Katayama, Y and Wood, A (1999). A re- evaluation of the of and a proposal for the combination Paracoccus pantotrophus comb. nov. International Journal of Systematic Bacteriology 49: 645-651.

Remold, S.K., Brown, C.K., Farris, J.E., Hundley, T.C., Perpich, J.A and Purdy, M.E (2011). Differential habitat use and niche partitioning by Pseudomonas species in human homes. Microbial Ecology 62: 505-517

Silyn-Roberts, G and Lewis, G (2001) In situ analysis of Nitrosomonas spp. in wastewater treatment wetland biofilms. Water Research 35, (11): 2731-2739

Spiers, A.J., Buckling, A and Rainey P.B (2000). The causes of Pseudomonas diversity. Microbiology 146 (10): 2345-2350

Yoon, J-H, Kang, S-S, Cho, Y-G., Lee, S.T., Kho, Y. H., Kim, C-J Park, Y-H (2000). Rhodococcus pyridinivorans sp. Nov., a pyridine-degrading bacterium. International Journal of Systematic and Evolutionary Microbiology 50: 2173-2180

4. Signature of applicant or person authorised to sign on behalf of applicant

X I request the Authority to waive any legislative information requirements (i.e. concerning the information that shall be supplied in my application) that my application does not meet (tick if applicable).

I have completed this application to the best of my ability and, as far as I am aware, the information I have provided in this application form is correct.

Signature Date

May 2012 EPA0159