DETERMINATION OF PATHOGEN RESISTANCE

TO STREPTOMYCIN

Lecturer Anca CHEREJI*, PhD

Rareş CHEREJI**, DVM

Abstract

This research was made in order to emphasize the actual incindence of the sensibility of various bacterial pathogens to streptomycin.
The pathogens were identified as belonging to Staphylococcus, Escherichia, Citrobacter, Enterobacter, Klebsiella, Salmonella, Proteus,
Pasteurella and Pseudomonas genus.

Antimicrobial agar disk diffusion method was used to test the strains isolated from animals and the results were interpreted as
resistant, intermediate and sensitive according to international standards.

Resistance in 100% percentage was registered for staphylococci and also for the pathogens from Citrobacter, Enterobacter,
Proteus, Pasteurella, Pseudomonas genus. There wasalso resistance for Escherichia coli strains - 90%, and Salmonella - 50%.

Intermediary values of the antibacterial inhibition zones presented Salmonella (50%) and also Escherichia coli pathogens (10%).
The only genus that was sensitive to streptomycin was Klebsiella.
The frequency of resistance phenomenon to streptomycin was 87,5%, intermediary values for the inhibition zones – 8,33% and

only 4,17% of the strains showed sensibility to this antibiotic.

Keywords: pathogen resistance, streptomycin

Introduction

Aminoglycosides represent, next to betalactamines, the group of antibiotics having the larger

therapeutical use. The first discovered aminoglycosides were natural molecules, produced by

Streptomyces (streptomycin, neomycin, kanamycin, tobramycin) or Micromonospora (gentamycin,

sisomycin).

Streptomycin and dihydrostreptomycin have a relatively restricted spectrum, the resistance being

very spread between these antibiotics. Nevertheless, there a few staphylococci, even some streptococci

and gram-negative bacilli still sensitive, among these Actinomyces bovis strains, Pasteurella spp, E. coli,

Salmonella spp., Campylobacter fetus, Leptospira spp. and Brucella spp.. Mycobacterium tuberculosis,

also, is still sensitive to streptomycin (Vakurenko S.B. and Mobashery S., 2003).

Starting with natural aminoglycosides, there were later conceived, using semisynthesis technique,

molecules less sensitive to enzyme inactivation induced by resistant bacteria and having lower toxicity

(amikacin, netilmycin etc.) (Oniga O. and Tiperciuc Brânduşa, 2003).

* University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca, Faculty of Veterinary Medicine, Department of
Pharmacology, 3-5 Mănăştur Street, 400372 Cluj-Napoca

** S.C. Biovet Serv. S.R.L., Cluj Napoca
Cluj Veterinary Journal, 15(1)/2009, pp. 69-72

This study was made in order to investigate the present resistance of some pathogens to streptomycin,

taking into account the fact that aminoglycosides are used, despite their high nephrotoxicitiy, to

control local and systemic infections due to sensitive aerobic bacteria (generally gram-negative)

(Cristina T. Romeo, 2006). The aminoglycosides recommendation is justified by their therapeutical

efficiency (Mărculescu Anca, 2007).

Material and Methods

The sensibility of a pathogen to antibiotics can be tested through a relative unsophisticated method

– the antibiogram.

The materials used in this study were represented by bacterial strains, cuture media and disks

with a standard quantity of antibiotic.

The bacterial strains were isolated from abscesses, phlegmons, vaginal secretions, secretions

mastitis, faeces, intestinal contents, organs with lesions (liver, spleen, heart, lungs, nodules etc.),

bones and corpses of animals.

The susceptibility to streptomycin was tested on 48 gram-positive and gram-negative bacterial

strains.

The cultivation was made on usual media (broth and agar) with a plus of serum, glucose, blood

when necessary, or special media (Răpuntean Gh. and Boldizsar E., 2001).

The interpretation of the antimicrobial agar disk diffusion method had been effected according

to international standards (NCCLS M31-A, M31-T, 1999).

Results and Discussions

A number of 48 pathogens from Staphylococcus, Escherichia, Citrobacter, Enterobacter, Klebsiella,

Salmonella, Proteus, Pasteurella, Pseudomonas genus was investigated through antibiogram in order

to determine the sensitivity to streptomycin.

The results were interpreted (table 1) and the pathogens were noted as resistant (R), sensible (S)

or with intermediary values (I) to this antibiotic (table 2).

Table1. Disc diffusion test interpretation according to international standards

Crt.
no.

Antibiotic
Atb/conc.

disc
Diametrul zonei de inhibiţie antibacteriană (mm)

R I S Standard values:

1
Streptomicină

Streptomicină

100 µg

10 µg

≤ 22

≤ 11

23 – 25

12 – 14

≥ 26

≥ 15

Neo-Sensitab Veterinary Pathogens –NCCLS

BBL Sensi-Disc – NCCLS

Table 2. Susceptibility/resistance to streptomycin

Crt.
no.

Bacterial strains No.
Sensibility/resistance

R % I % S %
1 Staphylococcus 6 6 100 - - - -
2 E. coli 20 18 90 2 10 - -
3 Citrobacter 2 2 100 - - - -
4 Enterobacter 2 2 100 - - - -
5 Klebsiella 2 - - - - 2 100
6 Salmonella 4 2 50 2 50 - -
7 Proteus 4 4 100 - - - -
8 Pasteurella 2 2 100 - - - -
9 Pseudomonas 6 6 100 - - - -

Total no. of pathogens 48 42 87,50 4 8,33 2 4,17

Many attainment mechanisms of resistance to streptomycin were described; can be plasmid mediated

or owed to mutations. A non-plasmid mediated mechanism is the decrease of the transportation

through cellular membrane. The anaerobic (Clostridium) and facultative anaerobic (enterobacteriaceae

and some staphylococci) bacteria are more resistant to aminoglycosides, in an anaerobic medium,

because of the active and oxygen-dependent transportation.

A 100% resistance of staphylococci can be observed in the antibiograms of the present study. The

resistance due to decreased carriage can be induced by the exposal of bacterial strains to sublethal

concentrations of antibiotic. An example is the Pseudomonas aeruginosa resistance to streptomycin,

as it is obvious also in the tested strains of this research.

The decreasing of the binding of antibiotic to ribosomes is also a mechanism of Pseudomonas

resistance to streptomycin; that type of mechanism is met also in Escherichia coli strains. Furthermore,

the results of the present study show a 100% percentage of resistance in Pseudomonas, whilst in

the case of Escherichia coli – a 90% resistance and 10% intermediary values of the antibacterial

inhibition zones; as some authors point out (Angelescu M., 1998), that means also 100% resistance,

because mostly statistical and epidemiological studies present the existence only of resistant and

sensible strains.

Even if Ray A. et al, in 2006, observe in Salmonella genus the highest resistance to streptomycin,

in this research there are only 50% of resistant strains; all the same, because of the existence of a great

number of intermediate strains – 50%, the sensitivity of the pathogens to this antibiotic is 0%.

The resistance emerge also through an enzy-

matic mechanism. These enzymes are found both

in gram-negative and gram-positive bacteria. It

is possible that this mechanism determined

the resistance in all strains from Citrobacter,

Enterobacter, Proteus şi Pasteurella genus, as it

is showed in graphic 1.

Accordingly, it is observed that for strepto-

mycin the antibiotic resistance phenomenon

appear in a rather high percentage – 87,5%,

as we marked for 21 of the 24 tested strains,

being in existence also a percentage of 8,33%

of intermediary values – 2 strains, and only one

strain was sensitive – a percentage of 4,17% of

the total strains (graphic 2).

Conclusions

Bacterial strains from Staphylococcus, Esche-

richia, Citrobacter, Enterobacter, Klebsiella,

Salmonella, Proteus, Pasteurella, Pseudomonas

genus were investigated about the susceptibility

to streptomycin;

The antimicrobial agar disk diffusion method was used for the testing of the pathogens;

The results were interpreted according to international standards;

Staphylococci registered a 100% resistance to streptomycin and also Citrobacter, Enterobacter,

Proteus, Pasteurella, Pseudomonas strains;

I % 8,33
S % 4,17

R % 87,5

100

0 20 40 60 80 100

Staf ilococi

E. coli

Citrobacter

Enterobacter

Klebsiella

Salmonella

Proteus

Pasteurella

Pseudomonas

S %
I %
R %

Graphic 1. Resistance to streptomycin

Graphic 2. The frequentness of resistance
phenomenon to streptomycin

Escherichia coli strains showed also rather high percentage of resistance to this antibiotic – 90%,

the rest of 10% being intermediary values of the inhibition zones

Resistance of 50% was observed for Salmonella pathogens and intermediary values, also in 50%

percentage

The only genus that was sensitive to streptomycin was Klebsiella;

The resistance of the investigated pathogens was, generally, 87,5%, being noted intermediary

values – 8,33% and only 4,17% of the strains showed sensibility to this antibiotic.

References

1. Angelescu Mircea – Terapia cu antibiotice, Ed. Medicală, Bucureşti, 1998;
2. Cristina T. Romeo – Bazele Farmacologiei Veterinare, Ed. Brumar, Timişoara, 2000;
3. Mărculescu Anca – PhD Thesis: „Studiu privind evoluţia fenomenului de antibiorezistenţă şi posibilitatea

diminuării acestuia prin asocierea de antibiotice, pe baza relaţiilor de sinergism”, Universitatea de
Ştiinţe Agricole şi Medicină Veterinară Cluj-Napoca, 2007

4. NCCLS, M31-A – Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for
Bacteria Isolated from Animals; Approved Standard, 1999, vol.19, No. 11;

5. NCCLS, M31-T – Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for
Bacteria Isolated from Animals; Approved Standard, 1999, vol.17, No. 11;

6. Oniga O., Brînduşa Tiperciuc – Antibiotice Antibacteriene, Editura Medicală Universitară „Iuliu
Haţieganu”, Cluj-Napoca, 2003;

7. Ray K.A., L.D. Warnick, R.M. Michell, J.B. Kaneene, P.L. Ruegg, S.J. Wells, C.P. Fossier, L.W. Halbert–
Antimicrobial susceptibility of Salmonella from organic and conventional daily farm, J. Dairy Sci.,
2006, 89 (6): 2038-50;

8. Răpuntean Gh., Boldizsar E. – Practicum de Bacteriologie Specială, Ed. AcademicPres, Cluj-Napoca,
2001;

9. Vakurenko S.B., S. Mobashery – Versatility of aminoglycosides and prospects for their future, Clinical
Microbiol. Reviews, 2003, vol. 16, 430-450;