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Leptotrichia goodfellowii is a fastidious, Gram-Negative bacillus belonging to the Fusobacteriaceae family. The bacterium is an aerotolerant anaerobe and is mesophilic, growing best at around the average human body temperature between 35℃-37℃, normal salinity, and a pH between 7.0-7.4 . Leptotrichia are a non-motile and non-spore forming bacterium that are a natural part of the microbiome of the genitourinary tract and oral cavities within humans and animals, however, L. goodfellowii is often only found in the blood in humans . They are also a rare cause of opportunistic infections, causing a variety of disease processes in immunocompromised persons.

Leptotrichia goodfellowii
Scientific classification
Kingdom:	Bacteria
Phylum:	Fusobacteria
Class:	Fusobacteriia
Order:	Fusobacteriales
Family:	Leptotrichiaceae
Genus:	Leptotrichia
Species:	goodfellowii
Binomial name
Leptotrichia goodfellowii

Taxonomy

Leptotrichia goodfellowii is one of seven known species that belong to the genus Leptotrichia. The first species discovered within this genus was L. buccalis. Other species include: L. hofstadii, L. honkongensis, L. shahii, L. trevisanii and L. wadei . Leptiotrichia was the first genus within the Leptotrichiaceae family, which also includes the genera Sebaldella, Sneathia, and Streptobacillus. Other members of the Leptotrichia genus have often been misclassified as other closely related genera. L. buccalis being most commonly misidentified as Fusobacterium or Lactobacillus. Original classification of L. goodfellowii was based upon 16S rRNA gene sequencing. L. goodfellowii had a 92% 16S rRNA gene sequence similarity to that of L. buccalis.

A recent study has proposed that L. goodfellowii be reclassified into a new genus, Pseudoleptotrichia. This reclassification has been proposed primarily due to L. goodfellowii being separate from the other six species when placed in a maximum-likelihood phylogenetic tree constructed using 16S rRNA sequences of each species. The six other Leptotrichia species share a 16S rRNA sequence similarity of 93.8-97.2%, while they are 88.7-90.6% similar to L. goodfellowii. L. goodfellowii, however, is 89.5% similar to the next related species of a different genus, Sebaldella termitidis. Furthermore, there are some differences in biochemical characteristics of L. goodfellowii. It is positive for β-Galactosidase and leucyl arylamidase which all other Leptotrichia species are negative for. It is negative for α-Glucosidase which the other six Leptotrichia species are positive for. With the combined molecular and biochemical differences, L. goodfellowii has been proposed to be reclassified as Pseudoleptotrichia goodfellowii in the year 2020.

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Discovery

L. goodfellowii was first discovered in 1683 by van Leeuwenhoek, who originally described the organism. The genus of Leptotrichia, established by Trevisan in 1879, was described as having a primary habitat of the human oral cavity, and producing lactic acid from fermenting carbohydrates from their metabolism. After the 16s rRNA gene was sequenced and a comparative analysis was done, Bergey’s Manual of 2005 placed the genus of Leptotrichia in the Fusobacteria phylum and the family of II Leptotrichiaceae. L. goodfellowii was named after Michael Goodfellow for his work in the field of microbial schematics . L. goodfellowii was isolated from human blood and colonies appeared on BHI agar plates after anaerobic incubation of 2-6 days. The BHI agar plates where the colonies grew were supplemented with 5% human blood, haemin, and menadione. The colonies grew scattered and irregular on the plate, looked pink on the outer ring and a light gray/brown color everywhere else, with a shiny surface, but dry and opaque inside. L. goodfellowii has been found in human oral cavities (as part of the normal flora), oral swabs of guinea pigs, and has been considered a human opportunistic pathogen.

Characterization

Leptotrichia goodfellowii is a fastidious, Gram-Negative bacillus belonging to the Fusobacteriaceae family, however, they can appear as gram-variable in their early growth stages due to their ability to hold on to crystal violet . The bacterium is an aerotolerant anaerobe and is mesophilic, growing best at around the average human body temperature between 35℃-37℃, normal salinity, and a pH between 7.0-7.4 . The bacterium is relatively small in comparison to other Leptotrichia species measuring around 2-4 μm long and 0.3-0.6 μm wide with a singular tapered end. The cells of Leptotrichia can be of either straight or curved rod shapes and tend to grow in pairs or chains that are joined by flattened ends. Leptotrichia are a non-motile and non-spore forming bacterium that are a natural part of the microbiome of the genitourinary tract and oral cavities within humans and animals, however, L. goodfellowii is often only found in the blood in humans . The bacterium is catalase-positive, oxidase-negative, beta-hemolytic, and an opportunistic pathogen linked to invasive diseases such as bacteremia and infective endocarditis . These systemic infections are rare and they primarily occur in immunocompromised individuals and individuals with mucosal lesions when the bacterium is translocated into damaged endothelial surfaces of blood vessels. It is difficult to clinically diagnose L. goodfellowii because Leptotrichia species can be easily misidentified as other species. The only way to accurately identify Leptotrichia goodfellowii is through molecular methods using 16s rRNA gene sequencing.

L. goodfellowii contains lipopolysaccharide (LPS) which displays an O-antigen that is linked to lipid-A. The Leptotrichia goodfellowii lipid-A endotoxin is often responsible for the toxic systemic symptoms, such as fever, shock, sepsis, and can also lead to spontaneous abortion. The bacterium’s LPS can trigger the host’s immune system into producing pro-inflammatory interleukins within epithelial cells in the body, which may lead to the harsh secondary effects of the infection.

Growth Conditions

L. goodfellowii is a fastidious bacteria, requiring serum and blood for growth, making it difficult to isolate from blood samples and grow in a culture . It grows best in anaerobic conditions and can grow in sparsely aerobic conditions, with 5-10% CO2, between a pH of 7.0-7.4, a normal salinity and a temperature of 35℃-37℃. It stops growing at temperatures 25℃ and below along with 42℃ and above. It can be grown on anaerobic blood agar for 2-6 days and shows beta-hemolytic patterns. They can also be grown on BHI or Colombia agar supplemented with 5% human blood, haemin, and menadione. The colonies formed appear convex, irregular, and would be pink in the outermost part and a greyish brown towards the center with a smooth surface. L. goodfellowii are susceptible to penicillins, vancomycin, and cephalosporins.

Metabolism

L. goodfellowii It is a chemoorganotroph and is obligately heterotrophic. It is able to ferment carbohydrates such as mannose, glucose, sucrose, lactose, and maltose to produce lactic acid. The bacterium can grow in the absence of a sugar-free medium, suggesting that fermentation of arginine can provide the ATP required for growth. It is the only species of Leptotrichia that is positive for the detection of beta-galactosidase and leucyl-arylamidase and negative for the detection of alpha-glucosidase.

Genomics

Whole genomes of twelve Leptotrichia species have been sequenced. The genome sequence of L. goodfellowii was originally assembled on January 27th, 2010 by Hans-Peter Klenk and Sabine Gronow. The complete genome on NCBI was sequenced using MinION and MiSeq and assembled via Canu v.1.8, Circlator v.1.5.5, nanopolish v.0.10.2. Pilon v.1.22 and CLC Genomics Workbench v.9.5.3. The genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP). PGAP combines alignment-based methods with methods that predict protein-coding and RNA genes. Species belonging to the Leptotrichia genus have been classified based upon 16S rRNA gene sequencing. The median genome length of L. goodfellowii is 2.28422 Mb with a median gene count of 2199 genes, protein count of 2079 proteins and a GC content of 31.55%. This is the highest GC content of all Leptotrichia species, which range from 29.3-31.55%.

Metabolic Potential

L. goodfellowii is incapable of growth on sucrose disaccharides due to premature stop codon at position 302 in the LepgoDRAFT_1710 position, causing a dysfunctional EII(CB) transporter of sucrose isomers. This organism has genetic loci for the phosphoenolpyruvate-dependent phosphotransferase systems for glucose and lactose. L. goodfellowii possesses 6 Type III toxin-antitoxin loci, which generally encode proteins that protect bacteria from infection by bacteriophages.

Ecology

Leptotrichia goodfellowii is normal member of the human microflora, specifically the oral, intestinal and female genital tracts. It is also an opportunistic pathogen that is typically isolated from the blood, but has also been collected from dog bites and guinea pig wounds. The ecology of this organism has primarily been determined via 16S rRNA gene sequencing and can be difficult to culture. Leptotrichia has not yet been isolated from environmental samples.

Significance

Members of the genus Leptotrichia are a common component of the oral, intestinal and genital microbiomes. However, they are also a rare cause of opportunistic infections, causing a variety of disease processes in immunocompromised persons. Examples of diseases caused by L. goodfellowii include infective endocarditis, cavities, aortic aneurysms, liver abscesses, and bacteremia. A common route of entry into the bloodstream for Leptotrichia is through oral mucosal injuries. A few case studies have shown L. goodfellowii to cause infective endocarditis in persons with well functioning immune systems, but a history of cardiac lesions. The exact prevalence of L. goodfellowii caused infections is unknown due to the difficulty in isolating and identifying members of the Leptotrichia genus. One cause of this difficulty is the number of Leptotrichia that retain crystal violet, causing them to be misidentified as Gram-positive rods, rather than Gram-negative rods. Common phenotypic and biochemical methods for identifying bacterial species are often ineffective for identifying L. goodfellowii. Retrospective studies using 16S rRNA gene sequencing have shown cases of L. goodfellowii that were previously missed by typical laboratory identification techniques. Several case studies have shown that L. goodfellowii can be treated with a wide variety of antibiotics and the only known incidences of resistance were to aminoglycosides, ciprofloxacin, and vancomycin.

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