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Acta Scientiarum 


ISSN printed: 1679-9283 
ISSN on-line: 1807-863X 
Doi: 10.4025/actascibiolsci.v39i3.34625 


Acta Scientiarum. Biological Sciences 

Maringá, v. 39, n. 3, p. 331-334, July-Sept., 2017 

Aerobic bacteria in oral cavity of Lancehead snakes (Bothrops atrox

with stomatitis  

Heloisa Castro Pereira


, Dayane Olímpia Gomes


, Líria Queiroz Luz Hirano


, André Luiz 

Quagliatto Santos


 and Anna Monteiro Correia Lima




Laboratório de Doenças Infecciosas, Faculdade de Medicina Veterinária, Universidade Federal de Uberlândia, Uberlândia, Minas Gerais, Brazil. 


Laboratório de Ensino e Pesquisa em Animais Silvestres, Faculdade de Medicina Veterinária, Universidade Federal de Uberlândia, Rua Piauí, 

s/n, 38400-902, Uberlândia, Minas Gerais, Brazil. *Author for correspondence. E-mail: liriaqueiroz@yahoo.com.br  

ABSTRACT. Stomatitis is a common disease found on snake farms, and Gram-negative bacilli are the 

main etiological agents that play an important role as secondary sources of viral or parasitic infections. The 
purpose of this work was to identify the aerobic bacteria in the oral cavity of Bothrops atrox with stomatitis. 
Samples for microbiological examination were collected from 12 snakes bred on a commercial snake farm 
for venom extraction. Samples of the secretion in the oral cavity of each serpent presenting stomatitis were 
collected from fang sheath, using a cotton swab with sterile alginate. The samples were incubated and 
cultured on Petri dishes containing blood agar and XLD agar using the agar depletion technique. Bacterial 
growth occurred in all analyzed samples collected from the oral cavity of Bothrops atrox with stomatitis, and 
some of the samples contained more than one microorganism. The following Gram-negative bacteria were 
isolated:  Escherichia coli (26.31%), Citrobacter spp. (21.05%), Proteus spp. (15.78%) and Salmonella spp. 
(10.52%). The only Gram-positive bacterium that was isolated was Staphylococcus spp., which was present in 
26.31% of the analyzed samples.


Keywords: bacteriology, infectious diseases, Reptilia, Squamata, Ophidia. 

Bactérias aeróbicas da cavidade oral de jararaca-do-norte (Bothrops atrox) com estomatite 

RESUMO. A estomatite é uma das doenças mais frequentes em criatórios comerciais de serpentes, sendo 
os bacilos Gram-negativos os principais agentes etiológicos com importante papel, como fontes 
secundárias, nas infecções virais ou parasitárias. O objetivo do presente trabalho foi analisar as bactérias 
aeróbicas presentes na cavidade oral em serpentes da espécie Bothrops atrox. Utilizaram-se 12 amostras 
colhidas com auxílio de swab estéril na região da bainha da presa, em serpentes que apresentaram 
estomatite, em um criatório comercial. As amostras foram cultivadas em Ágar-sangue e Ágar XLD. Em 
todas as amostras analisadas, houve crescimento de, pelo menos, um microrganismo. As bactérias Gram-
negativas isoladas foram Escherichia coli (26,31%), Citrobacter spp. (21,05%), Proteus spp. (15,78%) e Salmonella 
spp. (10,52%). A única bactéria Gram-positiva isolada foi a Staphylococcus spp., presente em 26,31% das 
amostras analisadas. 

Palavras-chave: bacteriologia, doenças infecciosas, Reptilia, Squamata, Ophidia. 


The gastrointestinal microbiota of reptiles 

generally comprises Gram-positive and Gram-

negative, aerobic and anaerobic yeasts and protozoa. 

Gram-negative bacilli are the main etiologic agents 

of diseases in snakes and play an important role as 

secondary sources of infections of viral or parasitic 

etiology. The prevalence of these bacteria in 

infectious processes is directly related to the 

opportunistic nature of the normal microbiota of 

reptiles (Cubas & Baptistotte, 2007).  

The stress that these reptiles normally undergoin 

the process of venom extraction and adaptation to an 

artificial environment has a direct impact on their 

health. This dynamic interferes with oral 

microbiota, which may, in turn, alter the amount 

and potency of venom (Soveri & Seuna, 1986). 

Bacterial diseases in snakes, which are 

characterized by different clinical manifestations, 
include stomatitis, ophthalmic and subcutaneous 
abscesses, glossitis, gastroenteritis, pneumonia, 
oophoritis and septicemia (Ferreira et al., 2012). 
Stomatitis is an important disease for animals used 
in venom obtention because of oral tissue injury and 
it was detected in 59% of snakes belonging to Boidae 
and Pythonidae families (Schmidt et al., 2013). 

Different authors have reported that the 

microorganisms commonly found as components of 

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Pereira et al. 

Acta Scientiarum. Biological Sciences 

Maringá, v. 39, n. 3, p. 331-334, July-Sept., 2017 

the microbiota of the digestive system of reptiles can 
act as etiological agents of gastrointestinal diseases, 

but few researchers have defined these bacteria in 

Brazilian species of reptiles. Moreover, the reports 

are scanty and many of them are outdated (Diaz-

Figueroa & Mitchell, 2006). 

Captive breeding of lancehead snakes has been 

extensive because the venom of these reptiles can be 

used in the production of drugs, which predisposes 

them to stomatitis. The objective of this study was 

to identify the aerobic bacteria in the oral cavity of 

captive Bothrops atrox suffering from stomatitis. 

Material and methods 

All protocols envolving animals have been 

reviewed and approved by SISBIO, Brazil’s 

Authorization and Information System on 

Biodiversity, under permit no. 41060-1, and the 

Universidade Federal de Uberlândia Ethics Committee 

(Animal use permite: 142/13). 

Samples for microbiological examination were 

collected from 12 snakes of the species Bothrops atrox 

which presented stomatitis, four males and eight 

females. The samples were taken from animals bred 
on a commercial snake farm for venom extraction 

belonging to the company Pentapharm do Brasil 

Comércio e Exportação LTDA, in Uberlândia, Minas 

Gerais, registered with the Brazilian Institute of 

Environment and Renewable Natural Resources 

(IBAMA) under no. 11904.  

To avoid adding to the animals’ stress, the 

samples were collected along with the routine 

procedures of the breeding farm, which follows all 

the animal welfare and biosafety standards. The 

samples were collected from snakes presenting 

stomatitis before they underwent any treatment. 

The snakes were physically restrained by placing 

a snake hook near the final third of the head, then 
positioning a hand in the region of the animal’s 
temporomandibular joints, thereby keeping its 
mouth open (Wilkinson, 2014). Samples of the 
secretion in the oral cavity of each serpent 
presenting stomatitis were then collected from its 
fang sheath, using a cotton swab


with sterile alginate 

(Figure 1) (Jorge et al., 1990). 

The samples were stored in tubes containing 

Stuart transport medium and sent to the Laboratory 
of Infectious Diseases of the Universidade Federal de 
. At the laboratory the samples were 
transferred to test tubes containing thioglycolate 
broth, a highly nutritious medium for culturing a 
variety of microorganisms, and were incubated in a 
bacteriological incubator at 37º C for 24 hours 
(Oplustil, 2004).  


Figure 1. Collecting material with a sterile swab in the region of 
the fang sheath. A. Healthy oral cavity of Bothrops atrox; B. Snake 
with stomatitis in the region of the right fang sheath. 

To isolate the bacterial colonies, the samples 

were cultured with the aid of a platinum wire loop 
on Petri dishes containing blood agar and XLDagar 
(Xylose Lysine Deoxycholate agar), using the agar 
depletion technique. The seeded dishes were then 
incubated again in a bacteriological incubator at  
37º C for 24 hours (Quinn et al., 2004). 

Colonies cultivated on blood agar were Gram 

stained to identify Gram-positive and Gram-
negative bacteria. Catalase and mannitol tests were 
used for the identification of Gram-positive bacteria 
(Oplustil, 2004). 

The colonies cultivated on XLDagar were 

identified using commercial Mini Kits containing 
Rugai medium with lysine, which is used mainly in 
biochemical screening of colonies grown on media 
selective for Gram-negative bacteria of the family 
Enterobacteriaceae. A fresh Rugai medium with 
lysine was employed separately for each different 
colony grown on XLDagar in order to identify each 
bacterial genus or species, as recommended by the 
manufacturer (Oplustil, 2004). 


Bacterial growth occurred in the all twelve 

analyzed samples collected from the oral cavity of 

Bothrops atrox with stomatitis, and some of the 
samples contained more than one microorganism. 

The following Gram-negative bacteria were isolated: 

Escherichia coli (26.31%),  Citrobacter  spp. (21.05%), 

Proteus  spp. (15.78%) and Salmonella  spp. (10.52%). 

The only Gram-positive bacterium that was isolated 

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Oral bacteria of Lancehead snakes 


Acta Scientiarum. Biological Sciences 

Maringá, v. 39, n. 3, p. 331-334, July-Sept., 2017 

was Staphylococcus spp., which was present in 26.31% 
of the analyzed samples (Table 1). 

Table 1. Frequency of bacteria found in the oral cavity of 

Bothrops atrox with stomatitis 


Number of 

positive samples 

Frequency (%) 

Escherichia coli 

5 26.31 

Staphylococcus spp. 


Citrobacter spp. 4 


Proteus spp. 3 


Salmonella spp. 2 




A study evaluated the aerobic microbiota in the 

oral cavity, cloaca and venom of specimens of the 

species  Crotalus durissus terrificus newly captured in 

the wild and quarantined, snakes kept in collective 

captivity, and snakes bred in individual captivity. 

The most common bacteria they found were 

Pseudomonas aeruginosaProteus vulgaris and Morganella 

morganii; however, Pseudomonas aeruginosa and 

Morganella morganii have not been found in Bothrops 

atrox (Ferreria Jr. et al., 2009). 

Snakes from different regions of the world 

present a variety of oral microbiota. Among the 

factors suggested to explain such differences are the 

particularities of the specimens evaluated, such as 

the species of snake, its origin, whether it is captive 

or free-living, its health status, whether or not it has 

fed recently, and the condition of the prey it feeds 

on (Jorge et al., 1990). 

The oral cavity of venomous and non-venomous 

snakes is colonized by a wide variety of anaerobic 

and aerobic microorganisms, and infections caused 

by  Aeromonas hydrophila,  Pseudomonas  spp., Proteus 

spp., Salmonella spp., Citrobacter spp., Escherichia coli, 

Providencia  spp., and Xanthomonas maltophilia stand 

out, among others (Blaylock, 2001). Gram-positive 

agents are also sometimes isolated, particularly 

Streptococcus  spp. and Staphylococcus  spp. (Cubas & 
Baptistotte, 2006). Staphylococcus  spp. has been 

identified frequently in Bothrops atrox, appearing in 

26% of samples. 

Some bacteria with a zoonotic potential have 

been isolated as part of the normal microbiota of 
snakes, and the main etiological agents are Gam-
negative bacilli such as the bacterium Morganella 
, which is distributed worldwide and is 
found in the normal microbiota of the oral and 
cloacal cavities of snakes. This microorganism is one 
of the bacterial agents involved in local 
complications by secondary necrosis in humans 
following a snake bite (Mader, 1998). In the analysis 
of a subcutaneous abscess in a Boa constrictor

Morganella morganii was the etiologic agent (Ferreira 
et al., 2012). 

Studies of Salmonella spp. in serpents suggest that 

the entry of infectious organisms in a snake 
collection is related not only to the addition of new 
reptiles but possibly also to the food that reptiles in 
captivity are fed, such as rodents (Williams, 2008). 

The prevalence of enterobacteria in Bothrops 

jararaca  in the state of São Paulo, based on a 
microbiological survey using samples taken directly 
from the colon of healthy adult snakes was 
evaluated. Several genera of the family 
Enterobacteriaceae were obtained (Citrobacter, 
Enterobacter, Escherichia, Klebsiella, Kluyvera, 
Morganella, Proteus, Providencia
 and Salmonella) and a 
morphologically similar genus of Gram-negative 
bacterium (Aeromonas).  Salmonella, Citrobacter and 
Escherichia  were the most frequent isolates. 
Escherichia, Proteus, Salmonella and  Citrobacter were 
also identified in the Bothrops atrox specimens 
analyzed (Bastos, Lopes, Gattamorta, & Matushima, 
2008). A study of the microbiota in the fangs, fangs 
heath and venom of Bothrops jararaca, reported that 
the bacterial species most frequently found were: 
group D streptococci, Enterobacter sp., Providencia 
, Providencia spp., Escherichia coli, Morganella 
and Clostridium sp., but less frequently 
Pseudomonas spp., Proteus mirabilis,  Staphylococcus 
,  Salmonella typhimurium, and Citrobacter  spp. 
(Jorge et al., 1990). 

A microbiological study of the oral cavity often 

different snake species belonging to the families 
Boidae, Colubridae, Elapidae and Viperidae, 
identified the bacteria Actinomyces  sp.,  Burkholderia 
spp.,  Moraxella  spp.,  Proteus  spp.,  Sarcina  spp., 
Bacillussubtilis,  Staphylococcus aureus, coagulase-
negative  Staphylococcus  and  Yersinia enterocolitica, and 
also isolated Proteus spp. and Staphylococcus spp. from 
Bothrops atrox (Fonseca, Moreira, Cunha, Ribeiro, & 
Almeida, 2009). 

Macroscopic and microscopic changes in 

Micrurus corallinus snakes kept in captivity, were 
observed in appearance of vesicles in the animals, 
and in their microbiological examination, isolated 
Staphylococcus aureus and  Pseudomonas aeruginosa
Blood cultures of these specimens were also 
performed using blood collected aseptically by 
intracardiac blood puncture, in which Staphylococcus 
was also isolated, indicating septicemia caused 
by this agent. Staphylococcus spp. was isolated in 26% 
of  Bothrops atrox samples analyzed (Serapicos et al., 

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Pereira et al. 

Acta Scientiarum. Biological Sciences 

Maringá, v. 39, n. 3, p. 331-334, July-Sept., 2017 


Bacterial growth was frequently present in the 

oral cavity of Bothrops atrox with stomatitis, and 

usually contain more than one species of 

microorganism. The most commonly isolated 

microorganisms were Gram-negative bacterium and 

the only Gram-positive bacterium that was isolated 

was Staphylococcus spp., which was present in 26.31% 

of the analyzed samples. 


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Received on December 21, 2016. 
Accepted on June 21, 2017. 



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