1.-­‐  MICROBIOLOGICAL  ANALYSIS  OF  THE  AIR  
 
Air,  in  addition  to  particles  in  suspension,  is  a  carrier  of  microorganisms  or  their  
spores.   In   addition   to   contributing   to   the   dispersion   of   microorganisms,   it   can  
cause  both  food  and  surface  contamination  and  infections  in  humans  and  animals  
(skin  and  respiratory  diseases).  
 
There   are   several   methods   for   air   analysis   such   as   sedimentation   or   filtration,  
among  others.  
 
 
1.1.-­‐  Plate  sedimentation  method  
 
It  is  one  of  the  simplest  methods  for  the  determination  of  air  microorganisms,  but  
the  results  are  indicative  of  the  numbers  of  microorganisms  present  in  it,  since  the  
values  obtained  depend  on  a  number  of  factors  such  as  air  currents  or  size  of  the  
particles  in  suspension.  
 
•  Leave  open  for  30,  60  and  90  minutes.  a  series  of  3  plates  with  general  medium  
TSA,  in  the  place  whose  level  of  contamination  we  want  to  study.  
•  Cover  plates  and  incubate  for  24-­‐48  hours  at  28oC.  
•   Observe   the   plates   and   isolate   the   colonies   that   have   appeared   for   later  
identification  in  selective  media.  
 
 
1.2.-­‐  Analysis  by  filtration  
 
To   do   this,   the   vacuum   pump   is   connected   and   aspirated   for   two   minutes.   The  
microorganisms  present  in  the  filtered  air  will  be  collected  in  the  membrane  and  
will   grow   when   it   is   deposited   on   the   surface   of   an   agar   plate   that   allows   the  
growth  of  the  aerobic  mesophilic  microorganisms.  The  bale  will  incubate  at  28oC  
for  24-­‐48  h.  
 
 
   
2.-­‐  MICROBIOLOGICAL  SURFACE  ANALYSIS  
 
Its   objective   is   to   check   the   hygienic   state   of   the   workplace.   There   are   several  
methods   available:   the   contact   plate   for   flat   surfaces   or   the   swab   for   non-­‐flat  
surfaces.  
 
2.1.-­‐   Analysis   of   flat   surfaces   by   means   of   contact   plates   (RODAC   replicate  
organisms  direct  agar  contact).  
 
•  Gently  press  an  RODAC  plate  on  the  chosen  surface  
•  Incubate  for  24-­‐48  h  at  37oC.  
•   Count   the   number   of   colonies   that   appeared   and   refer   them   to   the   sampled   area,  
9.6   cm2   (the   approximate   surface   area   on   the   surface   of   the   plate)   so   that   the  
result  will  be  expressed  as  a  CFU  /  9.6  cm2  number  
 
2.2.-­‐  Analysis  of  flat  and  non-­‐flat  surfaces:  method  of  the  swab.  
It  is  the  oldest  system  used  in  the  microbiological  analysis  of  surfaces,  especially  of  
appliances   and   utensils.   It   is   especially   recommended   to   study   highly  
contaminated  surfaces,  since  decimal  dilutions  of  the  sample  can  be  made.  
 
•  Delineate  the  surface  to  be  analyzed  using  a  sterile  aluminum  foil  template  of  9  
cm2.  
•   Moisten   the   sterile   swab   in   10   ml   of   sterile   saline   and   scrub   several   times   on   the  
surface  delimited  by  the  template.  
•   Put   the   swab   back   into   the   saline   tube   and   leave   for   15-­‐30   min   so   that   the  
microorganisms  are  released  from  the  cotton  to  the  liquid.  
•   Sow   0.1   ml   of   the   liquid   in   a   plate   of   TSA   (general   counting   medium)   and   /   or  
VRBG  (purple  violet  red  bile  glucose)  which  is  a  selective  and  differential  medium  
for  enterobacteria.  
•  Incubate  24-­‐48  h  at  37oC.  The  count  is  expressed  as  CFU  /  9cm2.  
 
 
3.-­‐  MICROBIOLOGICAL  SOIL  ANALYSIS  
 
The  soil  samples  are  taken  directly  from  it.  When  you  want  to  study  samples  of  a  
certain  depth,  you  must  have  a  device  (soil  corer)  that  facilitates  access  to  that  area  
and   that   when   entering   the   soil   does   not   agglomerate   it,   e.g.   a   hollow   tube   that  
penetrates  like  a  corkscrew  and  allows  the  control  of  its  filling.  
 
When  the  sampling  is  carried  out  in  a  soil  where  there  are  few  microorganisms,  an  
alternative  technique  can  be  made  that  allows  us  to  concentrate  them.  To  do  this,  a  
glass   slide   is   introduced   into   the   earth,   which   is   a   surface   to   which   the  
microorganisms   present   naturally   adhere.   In   this   way   a   representative   sample   of  
the  populations  of  that  soil  is  obtained.  
 
Alternatively,  you  can  introduce  a  capillary,  either  empty,  which  will  facilitate  the  
entry   of   microorganisms,   or   filled   with   a   nutrient   solution   that   will   then   serve   as   a  
means  of  enrichment,  so  it  will  not  serve  to  know  the  original  composition  of  the  
soil.  
 
 
3.1.-­‐  buried  slide  technique  
 
•   Fill   a   sterile   beaker   with   the   soil   to   be   studied   and   insert   a   sterile   slide   into   it.  
Leave  it  for  a  while  (2-­‐4  days).  
•  To  remove  the  slide,  tilt  it  lightly  towards  one  of  the  faces,  in  order  not  to  disturb  
the  microorganisms  adhered  to  the  upper  face.  
•  Tap  gently  on  the  table  to  remove  the  grains  of  soil.  
•  Clean  the  bottom  part  of  the  holder  with  a  filter  paper,  and  let  air  dry.