Phylogenetic analysis identifies the ‘megabacterium’ of birds as a novel anamorphic ascomycetous yeast, Macrorhabdus ornithogaster gen. nov., sp. nov.

Abstract

An organism commonly referred to as 'megabacterium' colonizes the gastric isthmus of many species of birds. It is weakly Gram-positive and periodic acid-Schiff-positive and stains with silver stains. Previous studies have shown that it has a nucleus and a cell wall similar to those seen in fungi. Calcofluor white M2R staining suggests that the cell wall contains chitin, a eukaryote-specific substance, and rRNA in situ hybridization demonstrates that it is a eukaryote. To characterize this organism phylogenetically, DNA was extracted from purified cells. rDNA was readily amplified by PCR with pan-fungal DNA primer sets and primer sets derived from the newly determined sequence, but not with bacteria-specific primer sets. Specific primer sets amplified rDNA from isthmus scrapings from an infected bird, but not from a non-infected bird or other control DNA. The sequence was confirmed to derive from the purified organism by in situ rRNA hybridization using a specific probe. Phylogenetic analysis of sequences of the 18S rDNA and domain D1/D2 of 26S rDNA showed the organism to be a previously undescribed anamorphic ascomycetous yeast representing a new genus. et al., 1980;Baker, 1992;Gerlach, 2001). These long, slender organisms (2-3620-80 mm) stain with silver stains and periodic acid-Schiff (PAS) and are weakly Gram-positive. The 'megabacterium' is found in the isthmus between the glandular and grinding stomach, where it grows on the lumenal surface and may penetrate koilin (Dorrestein et al., 1980;van Herck et al., 1984). It is associated with a lymphoplasmacytic gastritis in poultry (Mutlu et al., 1997) and a chronic fatal wasting disease in companion birds (Gerlach, 2001).The organism was originally thought to be a yeast because of its staining characteristics (Dorrestein et al., 1980). Subsequently, van Herck et al. (1984) concluded that it was a bacterium, as they were unable to demonstrate cytoplasmic organelles or a nucleus. They did, however, show nucleus-like structures in Geimsa-stained organisms, but interpreted them to be 'granules'. Scanlan & Graham (1990) reported isolating a bacterium from the stomach of a budgerigar using standard microbiological techniques. The isolated bacterium, however, was smaller than the organism observed in vivo and was not characterized by PAS or silver stains. Attempts by other investigators to grow this organism with standard microbiological isolation techniques have been unsuccessful. However, Gerlach (2001) reported isolation of this organism on MRS medium, but was unable to maintain it past a few passages. Huchzemeyer et al. (1993) also reported isolating an organism from the proventriculus of ostriches using MRS agar. This organism had the same biochemical properties as the one isolated byAbbreviations: D1/D2, domains 1 and 2; PAS, periodic acid-Schiff; PNA, peptide nucleic acid.

Analysis of fecal smears for diagnosis of megabactéria (Macrorhabdus ornithogaster) in domestic and wild birds

“…Apresenta sinais gastrointestinais graves como: diarreia, êmese, perda de peso progressiva, alimento mal digerido nas fezes, fezes aquosas e de cor verde-escura a marrom ou preta, e em casos mais graves fezes com sangue (WERTHER et al, 2000;DORRESTEIN, 2010;QUEIROS et al 2011). …”

“…Os sinais clínicos incluem também crescimento irregular, depressão, anorexia, disfagia, fraqueza, letargia, problemas com empenamento, além de dramática atrofia dos músculos peitorais, acúmulo de fezes ao redor da cloaca, queda de postura, baixo ganho de peso e caquexia (BAKER, 1997;WERTHER et al, 2000;MOORE et al, 2001).…”

“…236, Art. 1559, Julho, 2013 As amostras foram enviadas ao laboratório de análises clínicas da Universidade Guarulhos, onde foi realizado o esfregaço fecal, para pesquisa microscópica de megabactéria (Macrorhabdus ornithogaster) através da coloração de Gram e visualização em microscópio óptico comum com aumento de 1.000 vezes, assim como realizado nos estudos de Martins et al (2006) e Werther et al (2000.…”

“…A megabacteriose é uma enfermidade caracterizada por emagrecimento progressivo, regurgitação, alterações nas fezes, apatia e morte (WERTHER et al, 2000). Também chamada de "Síndrome Light Going", é causada por um fungo que afeta muitas espécies de aves, tendo sido associada a uma condição crônica sintomática ou assintomática (MOORE et al, 2001;GERLACH, 2001).…”

“…A afecção foi relatada em alguns países e em diversas espécies de aves, domésticas e silvestres, tais como: periquito australiano (Melopsittacus undulatus), canário (Serinus canarius), ema (Rhea americana), agapornis (Agapornis roseicollis), cacatua (Cacatua sp), calopsita ou gutarra (Nymphicus hollandicus), mandarim (Taeniopygia guttata), tucano (família Ramphastidae), avestruz (Struthio camelus), pomba (família Columbidae), peru (Meleagrins gallopavo) e codorna-japonesa (Coturnix coturnix) (WERTHER et al, 2000; QUEIROS, T.S., LOURENÇO, P. e PITA, M.C.G. Análise de esfregaço fecal para diagnóstico de megabactéria (Macrorhabdus ornithogaster) em aves domésticas e silvestres.…”